Aggregating microparticles for medical therapy

ABSTRACT

The present invention is a surface treated drug-loaded solid (e.g., non-porous) microparticle that aggregates in vivo to form a consolidated larger particle for medical therapy. In one embodiment, the particles are used for ocular therapy. Processes for producing the surface treated microparticle and injectable formulations which include the surface treated microparticle are also provided. When used in the eye, long-term consistent intraocular delivery can be achieved without disrupting vision and minimizing undesirable inflammatory responses.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.15/349,985, filed Nov. 11, 2016, which claims the benefit of U.S.Application No. 62/254,707, filed Nov. 12, 2015, U.S. Application No.62/257,608, filed Nov. 19, 2015, and U.S. Application No. 62/276,530,filed Jan. 8, 2016. The entirety of these applications is incorporatedby reference herein for all purposes.

FIELD OF THE INVENTION

The present invention is a surface treated drug-loaded solid (e.g.,non-porous) microparticle that aggregates in vivo to form a consolidatedlarger particle for medical therapy. In one embodiment, the particlesare used for ocular therapy. Processes for producing the surface treatedmicroparticle and injectable formulations, including the surface treatedmicroparticle, are also provided. When used in the eye, long-termconsistent intraocular delivery can be achieved that minimizesdisruption of vision and minimizes undesirable inflammatory responses.

BACKGROUND

The structure of the eye can be divided into two segments: the anteriorand posterior. The anterior segment comprises the front third of the eyeand includes the structures in front of the vitreous humor: the cornea,iris, ciliary body, and lens. The posterior segment includes the backtwo-thirds of the eye and includes the sclera, choroid, retinal pigmentepithelium, neural retina, optic nerve, and vitreous humor.

Important diseases affecting the anterior segment of the eye includeglaucoma, allergic conjunctivitis, anterior uveitis, and cataracts.Diseases affecting the posterior segment of the eye include dry and wetage-related macular degeneration (AMD), cytomegalovirus (CMV) infection,diabetic retinopathy, choroidal neovascularization, acute macularneuroretinopathy, macular edema (such as cystoid macular edema anddiabetic macular edema), Behcet's disease, retinal disorders, diabeticretinopathy (including proliferative diabetic retinopathy), retinalarterial occlusive disease, central retinal vein occlusion, uveiticretinal disease, retinal detachment, ocular trauma, damage caused byocular laser treatment or photodynamic therapy, photocoagulation,radiation retinopathy, epiretinal membrane disorders, branch retinalvein occlusion, anterior ischemic optic neuropathy, non-retinopathydiabetic retinal dysfunction and retinitis pigmentosa. Glaucoma issometimes also considered a posterior ocular condition because atherapeutic goal of glaucoma treatment is to prevent or reduce the lossof vision due to damage or loss of retinal cells or optic nerve cells.

Typical routes of drug administration to the eye include topical,systemic, intravitreal, intraocular, intracameral, subconjunctival,subtenon, retrobulbar, and posterior juxtascleral. (Gaudana, R., et al.,“Ocular Drug Delivery”, The American Association of PharmaceuticalScientist Journal, 12(3)348-360, 2010).

A number of types of delivery systems have been developed to delivertherapeutic agents to the eye. Such delivery systems includeconventional (solution, suspension, emulsion, ointment, inserts, andgels), vesicular (liposomes, niosomes, discomes, and pharmacosomes),advanced materials (scleral plugs, gene delivery, siRNA, and stemcells), and controlled-release systems (implants, hydrogels, dendrimers,iontophoresis, collagen shields, polymeric solutions, therapeuticcontact lenses, cyclodextrin carriers, microneedles, microemulsions, andparticulates (microparticles and nanoparticles)).

Treatment of posterior segment diseases remains a daunting challenge forformulation scientists. Drug delivery to the posterior segment of theeye is typically achieved via an intravitreal injection, the periocularroute, implant, or by systemic administration. Drug delivery to theposterior segment by way of the periocular route can involve theapplication of a drug solution to the close proximity of the sclera,which results in high retinal and vitreal concentrations.

Intravitreal injection is often carried out with a 30 gauge or lessneedle. While intravitreal injections offer high concentrations of drugto the vitreous chamber and retina, they can be associated with variousshort term complications such as retinal detachment, endophthalmitis andintravitreal hemorrhages. Experience shows that injection of smallparticles can lead to the rapid dispersal of the particles which canobstruct vision (experienced by the patient as “floaties” or “floaters”)and the rapid removal of the particles from the injection site (whichcan occur via the lymphatic drainage system or by phagocytosis). Inaddition, immunogenicity can occur upon recognition of the microspheresby macrophages and other cells and mediators of the immune system.

Complications in periocular injections include rises in intraocularpressure, cataract, hyphema, strabismus, and corneal decompensation.Transscleral delivery with periocular administration is seen as analternative to intravitreal injections. However, ocular barriers such asthe sclera, choroid, retinal pigment epithelium, lymphatic flow, andgeneral blood flow can compromise efficacy. Systemic administration,which is not advantageous given the ratio of the volume of the eye tothe entire body, can lead to potential systemic toxicity.

A number of companies have developed microparticles for treatment of eyedisorders. For example, Allergan has disclosed a biodegradablemicrosphere to deliver a therapeutic agent that is formulated in a highviscosity carrier suitable for intraocular injection or to treat anon-ocular disorder (U.S. publication 2010/0074957 and U.S. publication2015/0147406 claiming priority to a series of applications back to Dec.16, 2003). In one embodiment, the '957 application describes abiocompatible, intraocular drug delivery system that includes aplurality of biodegradable microspheres, a therapeutic agent, and aviscous carrier, wherein the carrier has a viscosity of at least about10 cps at a shear rate of 0.1/second at 25° C.

Allergan has also disclosed a composite drug delivery material that canbe injected into the eye of a patient that includes a plurality ofmicroparticles dispersed in a media, wherein the microparticles containa drug and a biodegradable or bioerodible coating and the media includesthe drug dispersed in a depot-forming material, wherein the mediacomposition may gel or solidify on injection into the eye (WO2013/112434 A1, claiming priority to Jan. 23, 2012). Allergan statesthat this invention can be used to provide a depot means to implant asolid sustained drug delivery system into the eye without an incision.In general, the depot on injection transforms to a material that has aviscosity that may be difficult or impossible to administer byinjection.

In addition, Allergan has disclosed biodegradable microspheres between40 and 200 μm in diameter, with a mean diameter between 60 and 150 μmthat are effectively retained in the anterior chamber of the eye withoutproducing hyperemia (US 2014/0294986). The microspheres contain a drugeffective for an ocular condition with greater than seven day releasefollowing administration to the anterior chamber of the eye. Theadministration of these large particles is intended to overcome thedisadvantages of injecting 1-30 μm particles which are generally poorlytolerated.

Regentec Limited has filed a series of patent applications on thepreparation of porous particles that can be used as tissue scaffolding(WO 2004/084968 and U.S. publication 2006/0263335 (filed Mar. 27, 2003)and U.S. publication 2008/0241248 (filed Sep. 20, 2005) and WO2008/041001 (filed Oct. 7, 2006)). The porosity of the particles must besufficient to receive cells to be held in the particle. The cells can beadded to the matrix at, or prior to, implantation of the matrix orafterward in the case of recruitment from endogenous cells in situ.Regentec also published an article on tissue scaffolding with porousparticles (Qutachi et al. “Injectable and porous PLGA microspheres thatform highly porous scaffolds at body temperature”, Acta Biomaterialia,10, 5080-5098, (2014)).

In addition, Regentec Limited also filed patent applications on thepreparation of large porous particles that can be used in drug delivery(WO 2010/100506 and U.S. publication 2012/0063997 (filed Mar. 5, 2009)).The porosity of the particles allows for quick delivery of thetherapeutic agent. The particles are intended to form a scaffold thatfills the space in which they are injected by a trigger such as a changein temperature.

Additional references pertaining to highly porous microparticles includepublications by Rahman and Kim. Rahman et al. “PLGA/PEG-hydrogelcomposite scaffolds with controllable mechanical properties” J. ofBiomedical Materials Research, 101, 648-655, (2013) describes hydrogelsof approximately 50 percent porosity and their corresponding mechanicalproperties. Kim et al. “Biodegradable polymeric microspheres with“open/closed” pores for sustained release of human growth hormone” J. ofControlled Release, 112, 167-174, (2006) describes PLGA polymers withpores for the delivery of human growth hormone.

EP 2125048 filed by Locate Therapeutics Limited (filed Feb. 1, 2007) aswell as WO 2008/093094, U.S. publication 2010/0063175 (filed Feb. 1,2007), and WO 2008/093095 (filed Feb. 1, 2007) filed by Regentec Limiteddisclose the preparation of particles that are not necessarily porousbut that when exposed to a trigger (such as temperature) form a tissuescaffold useful in the repair of damaged or missing tissue in a host.

U.S. Pat. No. 9,161,903 issued on Oct. 20, 2015 to Warsaw Orthopedic andU.S. publication 2016/0038407 filed by Warsaw Orthopedic Inc. disclose aflowable composition for injection at a target tissue site beneath theskin that includes a flowable composition that hardens at or near thetarget tissue site.

Bible et al. “Attachment of stem cells to scaffold particles forintra-cerebral transplantation”, Nat. Protoc., 10, 1440-1453, (2009)describes a detailed process to make microparticles of PLGA that do notclump or aggregate.

U.S. Patent Application Publication 2011/0123446 filed by LiquidiaTechnologies titled “Degradable compounds and methods of use thereof,particularly with particle replication in non-wetting templates”describes degradable polymers that utilize a silyl core and can formrapidly degrading matrixes.

Additional references pertaining to particles for ocular deliveryinclude the following. Ayalasomayajula, S. P. and Kompella, U. B. havedisclosed the subconjunctival administration of celecoxib-poly(lactideco-glycolide) (PLGA) microparticles in rats (Ayalasomayajula, S. P. andKompella, U. B., “Subconjunctivally administered celecoxib-PLGAmicroparticles sustain retinal drug levels and alleviatediabetes-induced oxidative stress in a rat model”, Eur. J. Pharm., 511,191-198 (2005)). Danbiosyst UK Ltd., has disclosed a microparticlecomprising a mixture of a biodegradable polymer, a water soluble polymerof 8,000 Daltons or higher and an active agent (U.S. Pat. No.5,869,103). Poly-Med, Inc. has disclosed compositions comprising ahydrogel mass and a carrier having a biological active agent depositedon the carrier (U.S. Pat. No. 6,413,539). MacroMed Inc. has disclosedthe use of an agent delivery system comprising a microparticle and abiodegradable gel (U.S. Pat. Nos. 6,287,588 and 6,589,549). Novartis hasdisclosed ophthalmic depot formulations for periocular orsubconjunctival administration where the pharmacologically acceptablepolymer is a polylactide-co-glycolide ester of a polyol (U.S.publication 2004/0234611, U.S. publication 2008/0305172, U.S.publication 2012/0269894, and U.S. publication 2013/0122064). TheUniversidad De Navarra has disclosed oral pegylated nanoparticles forcarrying biologically active molecules comprising a pegylatedbiodegradable polymer (U.S. Pat. No. 8,628,801). Surmodics, Inc. hasdisclosed microparticles containing matrices for drug delivery (U.S.Pat. No. 8,663,674). Minu, L.L.C., has disclosed the use of an agent inmicroparticle of nanoparticle form to facilitate transmembranetransport. Emory University and Georgia Tech Research Corporation havedisclosed particles dispersed in a non-Newtonian fluid that facilitatesthe migration of the therapeutic particles from the insertion site inthe suprachoroidal space to the treatment site (U.S. 2016/0310417).Pfizer has disclosed nanoparticles as injectable depot formulations(U.S. publication 2008/0166411). Abbott has disclosed a pharmaceuticaldosage form that comprises a pharmaceutically acceptable polymer for thedelivery of a tyrosine kinase inhibitor (U.S. publication 2009/0203709).The Brigham and Woman's Hospital, Inc. has disclosed modifiedpoly(lactic-co-glycolic) polymers having therapeutic agents covalentlybound to the polymer (U.S. 2012/0052041). BIND Therapeutics, Inc. hasdisclosed therapeutic nanoparticles comprising about 50 to 99.75 weightpercent of a diblock poly (lactic) acid-poly(ethylene)glycol copolymeror a diblock poly (lactic acid-co-glycolic acid)-poly(ethylene)glycolcopolymer wherein the therapeutic nanoparticle comprises 10 to about 30weight percent poly(ethylene)glycol (U.S. publication 2014/0178475).Additional publications assigned to BIND Therapeutics, Inc. include U.S.publication 2014/0248358 and U.S. publication 2014/0249158. Allergan hasdisclosed the use of biodegradable microspheres containing a drug totreat an ocular condition (U.S. publication 2010/0074957, U.S.publication 2014/0294986, U.S. publication 2015/0147406, EP 1742610, andWO 2013/112434). Allergan has also disclosed a biocompatible implantcontaining a prostamide component, which can exist in particle form, anda biodegradable polymer that allows for slow release of the drug overthe course of 1 week to 6 months for the treatment of an ocularcondition, such as glaucoma (U.S. application 2015/0157562 and U.S.application 2015/0099805). Jade Therapeutics has disclosed formulationscontaining an active agent and a polymer matrix that can be delivereddirectly to the target tissue or placed in a suitable delivery device(U.S. publication 2014/0107025). Bayer Healthcare has disclosed atopical ophthalmological pharmaceutical composition comprising sunitiniband at least one pharmaceutically acceptable vehicle (WO 2013/188283).pSivida Us, Inc. has disclosed biodegradable drug eluting particlescomprising a microporous or mesoporous silicon body for intraocular use(U.S. Pat. No. 9,023,896). Additional patents assigned to pSivida Us,Inc. include: U.S. Pat. Nos. 8,871,241; 8,815,284; 8,574,659; 8,574,613;8,252,307; 8,192,408 and 7,998,108. ForSight Vision4, Inc. has disclosedtherapeutic devices for implantation in the eye (U.S. Pat. No.8,808,727). Additional patents assigned to ForSight Vision4, Inc.include: U.S. Pat. Nos. 9,125,735; 9,107,748; 9,066,779, 9,050,765;9,033,911; 8,939,948; 9,905,963; 8,795,712; 8,715,346; 8,623,395;8,414,646; 8,399,006, 8,298,578; 8,277,830; 8,167,941; 7,883,520;7,828,844 and 7,585,075. The Nagoya Industrial Science ResearchInstitute has recently disclosed the use to liposomes to deliver a drugto the posterior segment of the eye (U.S. Pat. No. 9,114,070).

In order to treat ocular diseases, and in particular diseases of theposterior segment, the drug must be delivered in therapeutic levels andfor a sufficient duration to achieve efficacy. This seeminglystraightforward goal is difficult to achieve in practice.

The object of this invention is to provide compositions and methods totreat ocular disorders. Another objective is to provide drug deliveringmicroparticles for sustained administration of therapeutic materialsgenerally in vivo.

SUMMARY

The present invention provides mildly surface treated solidbiodegradable microparticles that on injection in vivo, aggregate to alarger particle (pellet) in a manner that reduces unwanted side effectsof the smaller particles and are suitable for long term (for example, upto, or alternatively at least, three months, four months, five months,six months or seven months or longer) sustained delivery of atherapeutic agent. In one embodiment, the mildly surface treated solidbiodegradable microparticles are suitable for ocular injection, at whichpoint the particles aggregate to form a pellet that remains outside thevisual axis so as not to significantly impair vision. The particles canaggregate into one or several pellets. The size of the aggregate dependson the concentration and volume of the microparticle suspensionsinjected and the diluent in which the microparticles are suspended.

In one embodiment, the invention is thus surface-modified solidaggregating microparticles that include at least one biodegradablepolymer, wherein the surface-modified solid aggregating microparticleshave a solid core, include a therapeutic agent, have a modified surfacewhich has been treated under mild conditions at a temperature at or lessthan about 18° C. to remove surface surfactant, are sufficiently smallto be injected in vivo, and are capable of aggregating in vivo to format least one pellet of at least 500 μm in vivo to provide sustained drugdelivery in vivo for at least one month, two months, three months, fourmonths, five months, six months or seven months or more. The surfacemodified solid aggregating microparticles are suitable, for example, foran intravitreal injection, implant, including an ocular implant,periocular delivery, or delivery in vivo outside of the eye.

In one embodiment, the surface-modified solid aggregating microparticlesdescribed herein, upon injection in vivo, aggregate in vivo to form atleast one pellet of at least 500 μm in vivo to provide sustained drugdelivery in vivo for at least one month, two months, three months, fourmonths, five months, six months or seven months or more.

In another embodiment, the invention is an injectable material thatincludes the microparticles of the present invention in apharmaceutically acceptable carrier for administration in vivo. Theinjectable material may include a compound that inhibits aggregation ofmicroparticles prior to injection and/or a viscosity enhancer and/or asalt. In one embodiment, the injectable material has a range ofconcentration of the surface-modified solid aggregating microparticlesof about 50 to 700 mg/ml. In certain examples, the injectable materialhas a concentration of the surface-modified solid aggregatingmicroparticles that is not more than about 50, 100, 150, 200, 250, 300,350, 400, 450, 500, 550, 600, 650 or 700 mg/ml. In one embodiment, theinjectable material has a concentration of the surface-modified solidaggregating microparticles of about 200-400 mg/ml, 150-450 or 100-500mg/ml. In certain embodiments, the injectable material has about up to150, 200, 300 or 400 mg/ml.

The present invention further includes a process for the preparation ofsurface-modified solid aggregating microparticles that includes

-   -   (i) a first step of preparing microparticles comprising one or        more biodegradable polymers by dissolving or dispersing the        polymer(s) and a therapeutic agent in one or more solvents to        form a polymer and therapeutic agent solution or dispersion,        mixing the polymer and the therapeutic agent solution or        dispersion with an aqueous phase containing a surfactant to        produce solvent-laden microparticles and then removing the        solvent(s) to produce polymer microparticles that contain the        therapeutic agent, polymer and surfactant; and    -   (ii) a second step of mildly treating the surface of        microparticles of step (i) at a temperature at or below about        18, 15, 10, 8 or 5° C. optionally up to about 1, 2, 3, 4, 5, 10,        30, 40, 50, 60, 70, 80, 90 100, 11, 120 or 140 minutes with an        agent that removes surface surfactant, surface polymer, or        surface oligomer in a manner that does not significantly produce        internal pores; and    -   (iii) isolating the surface treated microparticles.

The process can be achieved in a continuous manufacturing line or viaone step or in step-wise fashion. In one embodiment, wet biodegradablemicroparticles can be used without isolation to manufacture surfacetreated solid biodegradable microparticles. In one embodiment, thesurface treated solid biodegradable microparticles do not significantlyaggregate during the manufacturing process. In another embodiment, thesurface treated solid biodegradable microparticles do not significantlyaggregate when resuspended and loaded into a syringe. In someembodiments, the syringe is approximately 30, 29, 28, 27, 26 or 25gauge, with either normal or thin wall.

In yet another embodiment, a method for the treatment of an oculardisorder is provided that includes administering to a host in needthereof mildly surface-modified solid aggregating microparticles thatinclude an effective amount of a therapeutic agent, wherein thesurface-modified solid aggregating microparticles are injected into theeye and aggregate in vivo to form at least one pellet of at least 500 μmthat provides sustained drug delivery for at least approximately one,two, three, four, five, six or seven or more months in such a mannerthat the pellet stays substantially outside the visual axis so as not tosignificantly impair vision. In one embodiment, the surface treatedsolid biodegradable microparticles release about 1 to about 20 percent,about 1 to about 15 percent, about 1 to about 10 percent, or about 5 to20 percent, for example, up to about 1, 5, 10, 15 or 20 percent, of thetherapeutic agent over the first twenty-four (?) hour period. In oneembodiment, the surface treated solid biodegradable microparticlesrelease less therapeutic agent in vivo in comparison to non-treatedsolid biodegradable microparticles over up to about 1, 2, 3, 4, 5, 6, 7day or even up to about a 1, 2, 3, 4, or 5 month period. In oneembodiment, the surface treated solid biodegradable microparticlesinduce less inflammation in vivo in comparison to non-treated solidbiodegradable microparticles over the course of treatment.

This invention addresses the problem of intraocular therapy using smalldrug loaded particles (for example, 20 to 40 μm, 10 to 30, 20 to 30, or25 to 30 μm average diameter, or for example, not greater than about 20,25, 26, 27, 28, 29, 30, 35 or 40 μm average diameter (Dv)) that tend todisperse in the eye due to body movement and/or aqueous flow in thevitreous. The dispersed microparticles can cause vision disruption andaggravation from floaters, inflammation, etc. The microparticles of theinvention aggregate in vivo to form at least one pellet of at least 500μm and minimize vision disruption and inflammation. Further, theaggregated pellet of the surface treated microparticles is biodegradableso the aggregated pellet of the surface treated microparticles does nothave to be surgically removed.

In one embodiment, the surface treatment includes treatingmicroparticles with aqueous base, for example, sodium hydroxide and asolvent (such as an alcohol, for example ethanol or methanol, or anorganic solvent such as DMF, DMSO or ethyl acetate) as otherwisedescribed above. More generally, a hydroxide base is used, for example,potassium hydroxide. An organic base can also be used. In otherembodiments, the surface treatment as described above is carried out inaqueous acid, for example hydrochloric acid. In one embodiment, thesurface treatment includes treating microparticles with phosphatebuffered saline and ethanol.

In some embodiments, the surface treatment is carried out at atemperature of not more than 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16,17 or 18° C. at a reduced temperature of about 5 to about 18° C., about5 to about 16° C., about 5 to about 15° C., about 0 to about 10° C.,about 0 to about 8° C., or about 1 to about 5° C., about 5 to about 20°C., about 1 to about 10° C., about 0 to about 15° C., about 0 to about10° C., about 1 to about 8° C., or about 1 to about 5° C. Eachcombination of each of these conditions is considered independentlydisclosed as if each combination were separately listed.

The pH of the surface treatment will of course vary based on whether thetreatment is carried out in basic, neutral or acidic conditions. Whencarrying out the treatment in base, the pH may range from about 7.5 toabout 14, including not more than about 8, 9, 10, 11, 12, 13 or 14. Whencarrying out the treatment in acid, the pH may range from about 6.5 toabout 1, including not less than 1, 2, 3, 4, 5, or 6. When carrying outunder neutral conditions, the pH may typically range from about 6.4 or6.5 to about 7.4 or 7.5.

A key aspect of the present invention is that the treatment, whetherdone in basic, neutral or acidic conditions, includes a selection of thecombination of the time, temperature, pH agent and solvent that causes amild treatment that does not significantly damage the particle in amanner that forms pores, holes or channels. Each combination of each ofthese conditions is considered independently disclosed as if eachcombination were separately listed.

The treatment conditions should simply mildly treat the surface in amanner that allows the particles to remain as solid particles, beinjectable without undue aggregation or clumping, and form at least oneaggregate particle of at least 500 μm.

In one embodiment, the surface treatment includes treatingmicroparticles with an aqueous solution of pH=6.6 to 7.4 or 7.5 andethanol at a reduced temperature of about 1 to about 10° C., about 1 toabout 15° C., about 5 to about 15° C., or about 0 to about 5° C. In oneembodiment, the surface treatment includes treating microparticles withan aqueous solution of pH=6.6 to 7.4 or 7.5 and an organic solvent at areduced temperature of about 0 to about 10° C., about 5 to about 8° C.,or about 0 to about 5° C. In one embodiment, the surface treatmentincludes treating microparticles with an aqueous solution of pH=1 to 6.6and ethanol at a reduced temperature of about 0 to about 10° C., about 0to about 8° C., or about 0 to about 5° C. In one embodiment, the surfacetreatment includes treating microparticles with an organic solvent at areduced temperature of about 0 to about 18° C., about 0 to about 16° C.,about 0 to about 15° C., about 0 to about 10° C., about 0 to about 8°C., or about 0 to about 5° C. The decreased temperature of processing(less than room temperature, and typically less than 18° C.) assists toensure that the particles are only “mildly” surface treated.

Pharmaceutical and biologic therapeutic agents can be delivered in acontrolled fashion using the invention. In one embodiment, thepharmaceutical agent is a tyrosine kinase inhibitor such as sunitinib.One goal of the invention is to provide for the sustained release ofpharmaceutically active compounds to the eye, and in particular theposterior of the eye, over a period of at least about one, two, three,four, five, six, seven months or more in a manner that maintains atleast a concentration of a drug in the eye that is effective for thedisorder to be treated. In one embodiment, the drug is administered in asurface treated microparticle that provides for a sustained release thatis substantially linear. In another embodiment, the release is notlinear; however, even the lowest concentration of release over thedesignated time period is at or above a therapeutically effective dose.

In one embodiment, the surface treated microparticle includespoly(lactic-co-glycolic acid) (PLGA). In another embodiment, the surfacetreated microparticle includes a polymer or copolymer that has at leastPLGA and PLGA-polyethylene glycol (PEG) (referred to as PLGA-PEG). Inone embodiment, the surface treated microparticle includes poly(lacticacid) (PLA). In another embodiment, the surface treated microparticleincludes a polymer or copolymer that has at least PLA andPLA-polyethylene glycol (PEG) (referred to as PLA-PEG). In oneembodiment, the surface treated microparticle includes polycaprolactone(PCL). In another embodiment, the surface treated microparticle includesa polymer or copolymer that has at least PCL and PCL-polyethylene glycol(PEG) (referred to as PCL-PEG). In another embodiment, the surfacetreated microparticle includes at least PLGA, PLGA-PEG and polyvinylalcohol (PVA). In another embodiment, the surface treated microparticleincludes at least PLA, PLA-PEG and polyvinyl alcohol (PVA). In anotherembodiment, the surface treated microparticle includes at least PCL,PCL-PEG and polyvinyl alcohol (PVA). In other embodiments, anycombination of PLA, PLGA or PCL can be mixed with any combination ofPLA-PEG, PLGA-PEG or PCL-PEG, with or without PVA, and each combinationof each of these conditions is considered independently disclosed as ifeach were separately listed.

In one embodiment, the polyvinyl alcohol is a partially hydrolyzedpolyvinyl acetate. For example, the polyvinyl acetate is at least about78% hydrolyzed so that the polyvinyl acetate is substantiallyhydrolyzed. In one example, the polyvinyl acetate is at least about 88%to 98% hydrolyzed so that the polyvinyl acetate is substantiallyhydrolyzed.

In one embodiment, the surface treated microparticle including apharmaceutically active compound contains from about 80 percent or 89percent to about 99 percent PLGA, for example, at least about 80, 85,90, 95, 96, 97, 98 or 99 percent PLGA. In other embodiments, PLA or PCLis used in place of PLGA. In yet other embodiments, a combination ofPLA, PLGA and/or PCL is used.

In certain examples, the surface treated microparticle includes fromabout 0.5 percent to about 10 percent PLGA-PEG, about 0.5 percent toabout 5 percent PLGA-PEG, about 0.5 percent to about 4 percent PLGA-PEG,about 0.5 percent to about 3 percent PLGA-PEG, or about 0.1 percent toabout 1, 2, 5, or 10 percent PLGA-PEG. In other embodiments, PLA-PEG orPCL-PEG is used in place of PLGA-PEG. In other embodiments, anycombination of PLGA-PEG, PLA-PEG or PCL-PEG is used in the polymericcomposition with any combination of PLGA, PLA or PCL. Each combinationis considered specifically described as if set out individually herein.In one embodiment, the polymeric formulation includes up to about 1, 2,3, 4, 5, 6, 10, or 14% of the selected pegylated polymer.

In some examples, the microparticle contains from about 0.01 percent toabout 0.5 percent PVA (polyvinyl alcohol), about 0.05 percent to about0.5 percent PVA, about 0.1 percent to about 0.5 percent PVA, or about0.25 percent to about 0.5 percent PVA. In some examples, themicroparticle contains from about 0.001 percent to about 1 percent PVA,about 0.005 percent to about 1 percent PVA, about 0.075 percent to about1 percent PVA, or about 0.085 percent to about 1 percent PVA. In someexamples, the microparticle contains from about 0.01 percent to about5.0 percent PVA, about 0.05 percent to about 5.0 percent PVA, about 0.1percent to about 5.0 percent PVA, about 0.50 percent to about 5.0percent PVA. In some examples, the microparticle contains from about0.10 percent to about 1.0 percent PVA or about 0.50 percent to about 1.0percent. In some embodiments, the microparticle contains up to about0.10, 0.15, 0.20, 0.25, 0.30, 0.40 or 0.5% PVA. Any molecular weight PVAcan be used that achieves the desired results. In one embodiment, thePVA has a molecular weight of up to about 10, 15, 20, 25, 30, 35 or 40kd. In some embodiments, the PVA is partially hydrolyzed polyvinylacetate, including but not limited to, up to about 70, 75, 80, 85, 88,90 or even 95% hydrolyzed polyvinyl acetate. In one embodiment, the PVAis about 88% hydrolyzed polyvinyl acetate. In one embodiment, the PVApolymer has a molecule weight of 20,000 to 40,000 g/mol. In oneembodiment, the PVA polymer has a molecular weight of 24,000 to 35,000g/mol.

In one embodiment, the PLGA polymer has a molecular weight of 30,000 to60,000 g/mol (also kilodalton, kDa or kD). In one embodiment, the PLGApolymer has a molecular weight of 40,000 to 50,000 g/mol (for example40,000, 45,000 or 50,000 g/mol). In one embodiment, the PLA polymer hasa molecular weight of 30,000 to 60,000 g/mol (for example 40,000; 45,000or 50,000 g/mol). In one embodiment, the PCL polymer is used in the samerange of kDa as described for PLGA or PLA.

In one embodiment, a surface treated microparticle comprises apharmaceutically active compound. The encapsulation efficiency of thepharmaceutically active compound in the microparticle can range widelybased on specific microparticle formation conditions and the propertiesof the therapeutic agent, for example from about 20 percent to about 90percent, about 40 percent to about 85 percent, about 50 percent to about75 percent. In some embodiments, the encapsulation efficiency is forexample, up to about 50, 55, 60, 65, 70, 75 or 80 percent.

The amount of pharmaceutical active compound in the surface treatedmicroparticle is dependent on the molecular weight, potency, andpharmacokinetic properties of the pharmaceutical active compound.

In one embodiment, the pharmaceutically active compound is present in anamount of at least 1.0 weight percent to about 40 weight percent basedon the total weight of the surface treated microparticle. In someembodiments, the pharmaceutically active compound is present in anamount of at least 1.0 weight percent to about 35 weight percent, atleast 1.0 weight percent to about 30 weight percent, at least 1.0 weightpercent to about 25 weight percent, or at least 1.0 weight percent toabout 20 weight percent based on the total weight of the surface treatedmicroparticle. Nonlimiting examples of weight of active material in themicroparticle are at least about 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 or15% by weight. In one example, the microparticle has about 10% by weightof active compound.

In one embodiment, the invention provides a process for producing amicroparticle comprising a microparticle and a pharmaceutically activecompound encapsulated in the microparticle; which process comprises:

-   -   (a) preparing a solution or suspension (organic phase)        comprising: (i) PLGA or PLA (ii) PLGA-PEG or PLA-PEG (iii) a        pharmaceutically active compound and (iv) one or more organic        solvents;    -   (b) preparing an emulsion in an aqueous polyvinyl alcohol (PVA)        solution (aqueous phase) by adding the organic phase into the        aqueous phase and mixing at about 3,000 to about 10,000 rpm for        about 1 to about 30 minutes;    -   (c) hardening the emulsion including solvent-laden        microparticles including the pharmaceutically active compound by        stirring at about room temperature until solvent substantially        evaporates;    -   (d) centrifuging the microparticle including a pharmaceutically        active compound; (e) removing the solvent and washing the        microparticle including the pharmaceutically active compound        with water;    -   (f) filtering the microparticle including the pharmaceutically        active compound to remove aggregates or particles larger than        the desired size;    -   (g) optionally lyophilizing the microparticle comprising the        pharmaceutically active compound and storing the microparticle        as a dry powder in a manner that maintains stability for up to        about 6, 8, 10, 12, 20, 22, or 24 months or more.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 illustrates the aggregation of non-surface treated microparticles(NSTMP) (S-1 and S-5) and surface treated microparticles (STMP) (S-3 andS-8) after injection into PBS and incubation at 37° C. for 2 hours. TheNSTMP, S-1 and S-5, started to disperse immediately when the tubes wereinverted after the 2 hour-incubation, while the STMP, S-3 and S-8,remained aggregated at the bottom of the tubes without dispersionthroughout the entire period of observation (about 10 minutes). Samplesfrom left to right are S-1, S-3, S-5 and S-8 (Example 5).

FIG. 2 illustrates the aggregation of surface treated microparticles(STMP) (S-3 and S-8) after injection into HA and incubation at 37° C.for 2 hours. Samples left to right are S-1, S-3, S-5 and S-8 (Example5).

FIG. 3 illustrates the result of in vitro aggregation and dispersion ofparticles after a 2-hour incubation in PBS at 37° C. followed byagitation to detach the aggregates from the bottom of the tubes. Top rowfrom left to right samples: S-1, S-2, S-3, S-4; Bottom row from left toright samples: S-5, S-6, S-7 and S-8 (Example 5).

FIG. 4 illustrates in vitro aggregation of representative surfacetreated microparticles (STMP) treated with PBS/EtOH (sample S-21) aftera 2 hour incubation in PBS at 37° C. followed by agitation by tappingand flicking the tube (Example 6).

FIG. 5 illustrates the in vitro accelerated drug release profile of arepresentative batch of surface treated microparticles (STMP) (S-12)(Example 12). The x-axis is time measured in days and the y-axis iscumulative release percent.

FIG. 6 illustrates the in vitro drug release profiles for samples S-1,S-2, and S-3 in PBS with 1% Tween 20 at 37° C. (Example 13). The x-axisis time measured in days and the y-axis is cumulative release percent.

FIG. 7 illustrates the in vitro drug release profile of S-13, S-14, S-15and S-16 in PBS with 1% Tween 20 at 37° C. (Example 15). The x-axis istime measured in days and the y-axis is cumulative release percent.

FIG. 8A illustrates the in vitro aggregation of surface treatedmicroparticles (STMP) in 5-fold diluted ProVisc at a concentration of100 mg/mL into 4 mL of PBS after incubation at 37° C. for 2 hours (top)and after incubation at 37° C. for 2 hours followed by shaking at 250rpm for 2 minutes on an orbital shaker (bottom) (Example 17).

FIG. 8B illustrates the in vitro aggregation of surface treatedmicroparticles (STMP) in 5-fold diluted ProVisc at a concentration of100 mg/mL into 4 mL of HA (5 mg/mL solution) after incubation at 37° C.for 2 hours (top) and after incubation at 37° C. for 2 hours followed byshaking at 250 rpm for 2 minutes on an orbital shaker (bottom) (Example17).

FIG. 8C illustrates the in vitro aggregation of surface treatedmicroparticles (STMP) in 5-fold diluted ProVisc at a concentration of200 mg/mL into 4 mL of PBS after incubation at 37° C. for 2 hoursfollowed by shaking at 250 rpm for 2 minutes on an orbital shaker(bottom) (Example 17).

FIG. 8D illustrates the in vitro aggregation of surface treatedmicroparticles (STMP) in 5-fold diluted ProVisc at a concentration of200 mg/mL into 4 mL of HA (5 mg/mL solution) after incubation at 37° C.for 2 hours followed by shaking at 250 rpm for 2 minutes on an orbitalshaker (bottom) (Example 17).

FIG. 9 illustrates photos of aggregates of particles in an ex vivo coweye 2 hours after injection (Example 18).

FIG. 10A are photos of particle aggregates in the vitreous (left) andout of the vitreous (right) following injection of STMP, S-10, suspendedin PBS into the central vitreous of rabbit eyes (Example 19).

FIG. 10B are photos of particle aggregates in the vitreous (left) andout of the vitreous (right) following injection of STMP, S-10, suspendedin 5-fold diluted ProVisc into the central vitreous of rabbit eyes(Example 19).

FIG. 11A illustrates representative 1-month histology images of rabbiteyes injected with surface treated microparticles (STMP) (Example 20).

FIG. 11B illustrates representative 1-month histology images of rabbiteyes injected with non-surface treated microparticles (NSTMP) (Example20).

FIG. 12 illustrates the size distribution of a representative batch ofsurface treated microparticles (STMP) (S-12) (Example 22). The x-axisrepresents particle diameter measured in micrometers and the y-axisrepresents volume percent.

FIG. 13A illustrates select PK profiles for sunitinib in the retinafollowing a bilateral injection of sunitinib malate (free drug) at adose of 0.0125 mg/eye or 0.00125 mg/eye in pigmented rabbits (Example24). The x-axis is time measured in hours and the y-axis is theconcentration of sunitinib in ng/g.

FIG. 13B illustrates select PK profiles for sunitinib in the vitreousfollowing a bilateral injection of sunitinib malate (free drug) at adose of 0.0125 mg/eye or 0.00125 mg/eye in pigmented rabbits (Example24). The x-axis is time measured in hours and the y-axis is theconcentration of sunitinib in ng/g.

FIG. 13C illustrates select PK profiles for sunitinib in the plasmafollowing a bilateral injection of sunitinib malate (free drug) at adose of 2.5 mg/eye, 0.25 mg/eye, or 0.025 mg/eye in pigmented rabbits(Example 24). The x-axis is time measured in hours and the y-axis is theconcentration of sunitinib in ng/g.

FIG. 14 illustrates subitinib levels (ng/g) in rabbits injected with 10mg of STMP containing 1 mg sunitinib for 7 months post-dose. The rabbitswere sacrificed at 4 months and sunitinib levels (ng/g) were determinedin the vitreous, retina, plasma, and RPE-Choroid. Sunitinib levels wereabove the K_(i) for sunitinib against VEGFR and PDGFR (Example 20). Thex-axis represents time post-dose in month and the y-axis represents theconcentration of sunitinib measured in ng/g.

FIG. 15 illustrates sunitinib levels (ng/g) in rabbits injected with 2mg of STMP containing 0.2 mg sunitinib (10% w/w STMP) for 4 monthspost-dose. The rabbits were sacrificed at 4 months and sunitinib levels(ng/g) were determined in the vitreous, retina, plasma, and RPE-Choroid.Sunitinib levels were above the K_(i) for sunitinib against VEGFR andPDGFR in the RPE-Choroid and retina (Example 20). The x-axis representstime post-dose in months and the y-axis represents the concentration ofsunitinib measured in ng/g.

FIG. 16 illustrates sunitinib levels (ng/g) in rabbits injected with 10mg of STMP containing 0.2 mg sunitinib (2% w/w STMP). The rabbits weresacrificed at 4 months and sunitinib levels (ng/g) were determined inthe vitreous, retina, plasma, and RPE-Choroid. Sunitinib levels wereabove the K_(i) for sunitinib against VEGFR and PDGFR in the RPE-Choroidand retina (Example 20). The x-axis represents time post-dose in monthand the y-axis represents the concentration of sunitinib measured inng/g.

FIG. 17 illustrates the aggregation of surface treated microparticles(STMP) (S-28 to S-37 and S-12) after injection into PBS and incubationat 37° C. for 2 hours. After the 2 hour-incubation, the non-surfacetreated microparticles (NSTMP), S-27, became dispersed when the testtube was placed on an orbital shaker at 400 rpm for 30 seconds, whilethe surface treated microparticles (STMP), S-28 to S-37 and S-12,remained aggregated under the same agitation condition. Samples fromleft to right, top row to bottom row are S-28, S-29, S-30, S-31, S-32,S-33, S-34, S-35, S-36, S-37, S-12 and S-27 (Example 10).

FIG. 18 illustrates the aggregation of surface treated microparticles(STMP) (S-39 to S-45) after injection into PBS and incubation at 37° C.for 2 hours. After the 2 hour-incubation, the non-surface treatedmicroparticles (NSTMP), S-38, became dispersed when the test tube wasplaced on an orbital shaker at 400 rpm for 30 seconds, while the surfacetreated microparticles (STMP), S-39 to S-45, remained aggregated underthe same agitation condition. Samples from left to right, top row tobottom row are S-39, S-40, S-41, S-42, S-43, S-44 and S-45 (Example 10).

FIG. 19 is a graph depicting PK after a single IVT injection of STMPcontaining 1 mg sunitinib malate in rabbits. The rabbits were sacrificedat 10 days and 3 months and sunitinib levels (ng/g) were determined inthe vitreous, retina, and RPE-Choroid. Sunitinib levels were above theK_(i) for sunitinib against VEGFR and PDGFR in the RPE-Choroid andretina (Example 29). The x-axis represents time post-dose in moths andthe y-axis represents the concentration of sunitinib measured in ng/g.

DETAILED DESCRIPTION I. Terminology

Although specific terms are employed herein, they are used in a genericand descriptive sense only and not for purposes of limitation. Unlessotherwise defined, all technical and scientific terms used herein havethe same meaning as commonly understood by one of ordinary skill in theart to which this presently described subject matter belongs.

Compounds are described using standard nomenclature. Unless definedotherwise, all technical and scientific terms used herein have the samemeaning as is commonly understood by one of skill in the art to whichthis invention belongs.

The terms “a” and “an” do not denote a limitation of quantity, butrather denote the presence of at least one of the referenced item.

Recitation of ranges of values are merely intended to serve as ashorthand method of referring individually to each separate valuefalling within the range, unless otherwise indicated herein, and eachseparate value is incorporated into the specification as if it wereindividually recited herein. The endpoints of all ranges are includedwithin the range and are independently combinable. All methods describedherein can be performed in a suitable order unless otherwise indicatedherein or otherwise clearly contradicted by context. The use ofexamples, or exemplary language (e.g., “such as”), is intended merely tobetter illustrate the invention and does not pose a limitation on thescope of the invention unless otherwise claimed.

The term “carrier” refers to a diluent, excipient, or vehicle.

A “dosage form” means a unit of administration of a composition thatincludes a surface treated microparticle and a therapeutically activecompound. Examples of dosage forms include injections, suspensions,liquids, emulsions, implants, particles, spheres, creams, ointments,inhalable forms, transdermal forms, buccal, sublingual, topical, gel,mucosal, and the like. A “dosage form” can also include, for example, asurface treated microparticle comprising a pharmaceutically activecompound in a carrier.

The term “microparticle” means a particle whose size is measured inmicrometers (μm). Typically, the microparticle has an average diameterof from about 1 μm to 100 μm. In some embodiments, the microparticle hasan average diameter of from about 1 μm to 60 μm, for instance from about1 μm to 40 μm; from about 10 μm to 40 μm; from about 20 μm to 40 μm;from about 25 μm to 40 μm; from about 20 μm to 35 μm. For example, themicroparticle may have an average diameter of from 20 μm to 40 μm. Asused herein, the term “microsphere” means a substantially sphericalmicroparticle.

A “patient” or “host” or “subject” is typically a human, however, may bemore generally a mammal. In an alternative embodiment it can refer to,for example, a cow, sheep, goat, horse, dog, cat, rabbit, rat, mouse,bird and the like.

The term “mild” or “mildly” when used to describe the surfacemodification of the microparticles means that the modification(typically the removal of surfactant from the surface, as opposed to theinner core, of the particle) is less severe, pronounced or extensivethan when carried out at room temperature with the otherwise sameconditions. In general, the surface modification of the solidmicroparticles of the present invention is carried out in a manner thatdoes not create significant channels or large pores that wouldsignificantly accelerate the degradation of the microparticle in vivo,yet serves to soften and decrease the hydrophilicity of the surface tofacilitate in vivo aggregation.

The term “solid” as used to characterize the mildly surface treatedmicroparticle means that the particle is substantially continuous inmaterial structure as opposed to heterogeneous with significant channelsand large pores that would undesirably shorten the time ofbiodegradation.

II. Mildly Surface Treated Aggregating Microparticles and Methods

The present invention provides mildly surface treated solidbiodegradable microparticles that on injection in vivo, aggregate to alarger particle (pellet) in a manner that reduces unwanted side effectsof the smaller particles and are suitable for long term (for example, upto or at least three month, up to four month, up to five month, up tosix months, up to seven months or longer) sustained delivery of atherapeutic agent. In one embodiment, the lightly surface treated solidbiodegradable microparticles are suitable for ocular injection, at whichpoint the particles aggregate to form a pellet and thus remains outsidethe visual axis as not to significantly impair vision. The particles canaggregate into one or several pellets. The size of the aggregate dependson the mass (weight) of the particles injected.

The mildly surface treated biodegradable microparticles provided hereinare distinguished from “scaffold” microparticles, which are used fortissue regrowth via pores that cells or tissue material can occupy. Incontrast, the present microparticles are designed to be solid materialsof sufficiently low porosity that they can aggregate to form a largercombined particle that erodes primarily by surface erosion for long termcontrolled drug delivery.

The surface modified solid aggregating microparticles of the presentinvention are suitable, for example, for intravitreal injection,implant, periocular delivery, or delivery in vivo outside the eye.

The surface modified solid aggregating microparticles of the presentinvention are also suitable for systemic, parenteral, transmembrane,transdermal, buccal, subcutaneous, endosinusial, intra-abdominal,intra-articular, intracartilaginous, intracerebral, intracoronal,dental, intradiscal, intramuscular, intratumor, topical, or vaginaldelivery in any manner useful for in vivo delivery.

In one embodiment, the invention is thus surface-modified solidaggregating microparticles that include at least one biodegradablepolymer, wherein the surface-modified solid aggregating microparticleshave a solid core, include a therapeutic agent, have a modified surfacewhich has been treated under mild conditions at a temperature at or lessthan about 18° C. to remove surface surfactant, are sufficiently smallto be injected in vivo, and aggregate in vivo to form at least onepellet of at least 500 μm in vivo in a manner that provides sustaineddrug delivery in vivo for at least one, two, three, four, five, six orseven months or more. The surface modified solid aggregatingmicroparticles are suitable, for example, for an intravitreal injection,implant, including an ocular implant, periocular delivery or delivery invivo outside of the eye.

Alternatively, the surface treatment is conducted at a temperature at orless than about 10° C., 8° C. or 5° C.

The surface treatment can be carried out at any pH that achieves thedesired purpose. Nonlimiting examples of the pH are between about 6 andabout 8, 6.5 and about 7.5, about 1 and about 4; about 4 and about 6;and 6 and about 8. In one embodiment the surface treatment can beconducted at a pH between about 8 and about 10. In one embodiment thesurface treatment can be conducted at a pH between about 10.0 and about13.0. In one embodiment the surface treatment can be conducted at a pHbetween about 12 and about 14. In one embodiment the surface treatmentcan be conducted with an organic solvent. In one embodiment the surfacetreatment can be conducted with ethanol. In other various embodiments,the surface treatment is carried out in a solvent selected frommethanol, ethyl acetate and ethanol. Nonlimiting examples are ethanolwith an aqueous organic base; ethanol and aqueous inorganic base;ethanol and sodium hydroxide; ethanol and potassium hydroxide; anaqueous acidic solution in ethanol; aqueous hydrochloric acid inethanol; and aqueous potassium chloride in ethanol.

Examples of solid cores included in the present invention include solidcores comprising a biodegradable polymer with less than 10 percentporosity, 8 percent porosity, 7 percent porosity, 6 percent porosity, 5percent porosity, 4 percent porosity, 3 percent porosity, or 2 percentporosity. Porosity as used herein is defined by ratio of void space tototal volume of the surface-modified solid aggregating microparticle.

The surface-modified solid aggregating microparticles of the presentinvention provides sustained delivery for at least one month, or atleast two months, or at least three months, or at least four months, orat least five months, or at least six months, or at least seven months.

The therapeutic agent delivered by the surface-modified solidaggregating microparticle is in one embodiment a pharmaceutical drug ora biologic. In nonlimiting examples, the pharmaceutical drugs includesunitinib, another tyrosine kinase inhibitor, an anti-inflammatory drug,an antibiotic, an immunosuppressing agent, an anti-VEGF agent, ananti-PDGF agent, or other therapeutic agents as described below.

In one embodiment the surface-modified solid aggregating microparticlehas a mean diameter between 10 and 60 μm, 20 and 50 μm, 20 and 40 μm, 20and 30 μm, 25 and 40 μm, or and 35 μm.

Further, the surface-modified solid aggregating microparticles of thedisclosed invention can aggregate to produce at least one pellet whenadministered in vivo that has a diameter of at least about 300, 400, 500μm, 600 μm, 700 μm, 1 mm, 1.5 mm, 2 mm, 3 mm, 4 mm, or 5 mm.

In one embodiment the surface-modified solid aggregating microparticlesof the present invention produces a pellet in vivo that releases thetherapeutic agent without a burst of more than about 10 percent or 15percent of total payload over a one week, or a five, four, three, twoday or one day period.

In some embodiments, the long term controlled drug delivery isaccomplished by a combination of surface erosion of an aggregatedmicroparticle over several months (for example, one, two, three, or fourmonths or more) followed by erosion of remaining parts of the aggregatedmicroparticle, followed by slow release of active material from in vivoproteins to which it has bound over the period of long term release fromthe aggregated particle. In another embodiment, the microparticledegrades substantially by surface erosion over a period of at leastabout one, two, three, four, five or six months or more.

In another embodiment the surface-modified solid aggregatingmicroparticles of the present invention have a drug loading of 1-40percent, 5-25 percent, or 5-15 percent weight/weight.

Examples of polymeric compositions included in surface-modified solidaggregating microparticles of the present invention include, but are notlimited to poly(lactide co-glycolide), poly(lactide-co-glycolide)covalently linked to polyethylene glycol, more than one biodegradablepolymer or copolymer mixed together, for example, a mixture ofpoly(lactide-co-glycolide) and poly(lactide-co-glycolide) covalentlylinked to polyethylene glycol, poly(lactic acid), a surfactant, such aspolyvinyl alcohol (which can be hydrolyzed polyvinyl acetate).

In another embodiment, the invention is an injectable material thatincludes the microparticles of the present invention in apharmaceutically acceptable carrier for administration in vivo. Theinjectable material may include a compound that inhibits aggregation ofmicroparticles prior to injection and/or a viscosity enhancer and/or asalt. In one embodiment, the injectable material has a range ofconcentration of the surface-modified solid aggregating microparticlesof about 50-700 mg/ml, 500 or less mg/ml, 400 or less mg/ml, 300 or lessmg/ml, 200 or less mg/ml, or 150 or less mg/ml.

The present invention further includes a process for the preparation ofsurface-modified solid aggregating microparticles that includes

-   -   (i) a first step of preparing microparticles comprising one or        more biodegradable polymers by dissolving or dispersing the        polymer(s) and a therapeutic agent in one or more solvents to        form a polymer and therapeutic agent solution or dispersion,        mixing the polymer and the therapeutic agent solution or        dispersion with an aqueous phase containing a surfactant to        produce solvent-laden microparticles and then removing the        solvent(s) to produce microparticles that contain the        therapeutic agent, polymer and surfactant: and    -   (ii) a second step of mildly surface-only treating the        microparticles of step (i) at a temperature at or below about        18° C. for optionally up to about 140, 120, 110, 100, 90, 80,        70, 60, 50, 40, 30, 10, 8, 5, 4, 3, 2, or 1 minutes with an        agent that removes surface surfactant, surface polymers, or        surface oligomers in a manner that does not significantly        produce internal pores; and    -   (iii) isolating the surface treated microparticles.

In certain embodiments step (ii) above is carried out at a temperaturebelow 17° C., 15° C., 10° C., or 5° C. Further, step (iii) is optionallycarried out at a temperature below 25° C., below 17° C., 15° C., 10° C.,8° C. or 5° C. Step (ii), for example, can be carried out for less than8, less than 6, less than 4, less than 3, less than 2, or less than 1minutes. In one embodiment, step (ii) is carried out for less than 60,50, 40, 30, 20, or 10 minutes.

In one embodiment, the process of manufacturing surface-modified solidaggregating microparticles includes using an agent that removes surfacesurfactant. Nonlimiting examples include for example, those selectedfrom: aqueous acid, phosphate buffered saline, water, aqueous NaOH,aqueous hydrochloric acid, aqueous potassium chloride, alcohol orethanol.

In one embodiment, the process of manufacturing surface-modified solidaggregating microparticles includes using an agent that removes surfacesurfactant which comprises, for example, a solvent selected from analcohol, for example, ethanol; ether, acetone, acetonitrile, DMSO, DMF,THF, dimethylacetamide, carbon disulfide, chloroform,1,1-dichloroethane, dichloromethane, ethyl acetate, heptane, hexane,methanol, methyl acetate, methyl t-butyl ether (MTBE), pentane,propanol, 2-propanol, toluene, N-methyl pyrrolidinone (NMP), acetamide,piperazine, triethylenediamine, diols, and CO₂.

The agent that removes the surface surfactant can comprise a basicbuffer solution. Further, the agent that removes surface surfactant cancomprises a base selected from sodium hydroxide, lithium hydroxide,potassium hydroxide, calcium hydroxide, magnesium hydroxide, lithiumamide, sodium amide, barium carbonate, barium hydroxide, bariumhydroxide hydrate, calcium carbonate, cesium carbonate, cesiumhydroxide, lithium carbonate, magnesium carbonate, potassium carbonate,sodium carbonate, strontium carbonate, ammonia, methylamine, ethylamine,propylamine, isopropylamine, dimethylamine, diethylamine, dipropylamine,diisopropylamine, trimethylamine, triethylamine, tripropylamine,triisopropylamine, aniline, methylaniline, dimethylaniline, pyridine,azajulolidine, benzylamine, methylbenzylamine, dimethylbenzylamine,DABCO, 1,5-diazabicyclo[4.3.0]non-5-ene,1,8-diazabicyclo[5.4.0]non-7-ene, 2,6-lutidine, morpholine, piperidine,piperazine, Proton-sponge, 1,5,7-Triazabicyclo[4.4.0]dec-5-ene,tripelennamine, ammonium hydroxide, triethanolamine, ethanolamine, andTrizma.

In one embodiment, the process of manufacturing surface-modified solidaggregating microparticles includes using an agent that removes surfacesurfactant, for example, those selected from the following: aqueousacid, phosphate buffered saline, water, or NaOH in the presence of asolvent such as an alcohol, for example, ethanol, ether, acetone,acetonitrile, DMSO, DMF, THF, dimethylacetamide, carbon disulfide,chloroform, 1,1-dichloroethane, dichloromethane, ethyl acetate, heptane,hexane, methanol, methyl acetate, methyl t-butyl ether (MTBE), pentane,ethanol, propanol, 2-propanol, toluene, N-methyl pyrrolidinone (NMP),acetamide, piperazine, triethylenediamine, diols, and CO₂.

In one embodiment, the agent that removes the surface surfactant cancomprise an aqueous acid. The agent that removes the surface surfactantcan comprise an acid derived from inorganic acids including, but notlimited to, hydrochloric, hydrobromic, sulfuric, sulfamic, phosphoric,nitric and the like; or organic acids including, but not limited to,acetic, propionic, succinic, glycolic, stearic, lactic, malic, tartaric,citric, ascorbic, pamoic, maleic, hydroxymaleic, phenylacetic, glutamic,benzoic, salicylic, mesylic, esylic, besylic, sulfanilic,2-acetoxybenzoic, fumaric, toluenesulfonic, methanesulfonic, ethanedisulfonic, oxalic, isethionic, HOOC—(CH₂)_(n)—COOH where n is 0-4, andthe like.

In one embodiment, the agent that removes surface surfactant is not adegrading agent of the biodegradable polymer under the conditions of thereaction. The hydrophilicity of the microparticles can be decreased byremoving surfactant.

In one embodiment, the process of manufacturing surface-modified solidaggregating microparticles comprises using an agent that removes surfacesurfactant that comprises a solvent selected from an alcohol, forexample, ethanol, ether, acetone, acetonitrile, DMSO, DMF, THF,dimethylacetamide, carbon disulfide, chloroform, 1,1-dichloroethane,dichloromethane, ethyl acetate, heptane, hexane, methanol, methylacetate, methyl t-butyl ether (MTBE), pentane, ethanol, propanol,2-propanol, toluene, N-methyl pyrrolidinone (NMP), acetamide,piperazine, triethylenediamine, diols, and CO₂. In a typical embodimentthe process of surface treating, comprises an agent that removes surfacesurfactant that comprises ethanol.

The encapsulation efficiency of the process of manufacturingsurface-modified solid aggregating microparticles depends on themicroparticle forming conditions and the properties of the therapeuticagent. In certain embodiments, the encapsulation efficiency can begreater than about 50 percent, greater than about 75 percent, greaterthan about 80 percent, or greater than about 90 percent.

In one embodiment, the process of manufacturing surface-modified solidaggregating microparticles includes 75/25 PLGA as a biodegradablepolymer.

In an alternative embodiment, the process of manufacturingsurface-modified solid aggregating microparticles is carried out belowabout a pH of 14 and above a pH of 12, below a pH of 12 and above a pHof 10, below a pH of about 10 and above a pH of 8, below about a pH of 8and above a pH of about 6, neutral pH, below about a pH of 7 and above apH of 4, below about a pH of 4 and above a pH of about 1.0.

In one embodiment, step (ii) above is carried out for a time of aboutless than 140, 120, 110, 100, 90, 60, 40, 30, 20, 10, 3, 2, or 1minutes.

In yet another embodiment, a method for the treatment of an oculardisorder is provided that includes administering to a host in needthereof surface-modified solid aggregating microparticles that includean effective amount of a therapeutic agent, wherein the therapeuticagent containing surface-modified solid aggregating microparticles areinjected into the eye and its vivo aggregate to form at least one pelletof at least 500 μm that provides sustained drug delivery for at leastone, two, or three, four, five, six, seven or more months in such amanner that the pellet stays substantially outside the visual axis asnot to significantly impair vision.

In an alternative embodiment, the weight percent of surface-modifiedsolid aggregating microparticles that are not aggregated into a largerpellet in vivo is about 10 percent or less, 7 percent or less, 5 percentor less, or 2 percent or less by total weight administered.

In one embodiment, the surface-modified solid aggregating microparticlesdo not cause substantial inflammation in the eye.

In another embodiment, the surface-modified solid aggregatingmicroparticles do not cause an immune response in the eye.

In one embodiment, the surface-modified microparticles of the presentinvention, as described herein, are used to treat a medical disorderwhich is glaucoma, a disorder mediated by carbonic anhydrase, a disorderor abnormality related to an increase in intraocular pressure (IOP), adisorder mediated by nitric oxide synthase (NOS), or a disorderrequiring neuroprotection such as to regenerate/repair optic nerves. Inanother embodiment more generally, the disorder treated is allergicconjunctivitis, anterior uveitis, cataracts, dry or wet age-relatedmacular degeneration (AMD), or diabetic retinopathy.

Another embodiment is provided that includes the administration of asurface treated microparticle comprising an effective amount of apharmaceutically active compound or a pharmaceutically acceptable saltthereof, optionally in a pharmaceutically acceptable carrier, to a hostto treat an ocular or other disorder that can benefit from topical orlocal delivery. The therapy can be delivery to the anterior or posteriorchamber of the eye. In specific aspects, a surface treated microparticlecomprising an effective amount of a pharmaceutically active compound isadministered to treat a disorder of the cornea, conjunctiva, aqueoushumor, iris, ciliary body, lens sclera, choroid, retinal pigmentepithelium, neural retina, optic nerve, or vitreous humor.

Any of the compositions described can be administered to the eye asdescribed further herein in any desired form of administration,including via intravitreal, intrastromal, intracameral, subtenon,sub-retinal, retrobulbar, peribulbar, suprachoroidal, subchoroidal,conjunctival, subconjunctival, episcleral, posterior juxtascleral,circumcorneal, tear duct injections, or through a mucus, mucin, or amucosal barrier, in an immediate or controlled release fashion.

In one embodiment the disclosure provides a beta-adrenergic antagonistfor ocular therapy, which can be released from a surface treatedmicroparticle while maintaining efficacy over an extended time such asup to at least 1, 2, 3, 4, 5, 6, or 7 months.

In one embodiment the disclosure provides a prostaglandin analog forocular therapy, which can be released from a surface treatedmicroparticle while maintaining efficacy over an extended time such asup to at least 1, 2, 3, 4, 5, 6, or 7 months.

In one embodiment the disclosure provides an adrenergic agonist forocular therapy, which can be released from a surface treatedmicroparticle while maintaining efficacy over an extended time such asup to at least 1, 2, 3, 4, 5, 6, or 7 months.

In one embodiment the disclosure provides a carbonic anhydrase inhibitorfor ocular therapy, which can be released from a surface treatedmicroparticle while maintaining efficacy over an extended time such asup to at least 1, 2, 3, 4, 5, 6, or 7 months.

In one embodiment the disclosure provides a parasympathomimetic agentfor ocular therapy, which can be released from a surface treatedmicroparticle while maintaining efficacy over an extended time such asup to at least 1, 2, 3, 4, 5, 6, or 7 months.

In one embodiment the disclosure provides a dual anti-VEGF/anti-PDGFagent for ocular therapy, which can be released from a surface treatedmicroparticle while maintaining efficacy over an extended time such asup to at least 1, 2, 3, 4, 5, 6, or 7 months.

Methods of treating or preventing ocular disorders, including glaucoma,a disorder mediated by carbonic anhydrase, a disorder or abnormalityrelated to an increase in intraocular pressure (IOP), a disordermediated by nitric oxide synthase (NOS), a disorder requiringneuroprotection such as to regenerate/repair optic nerves, allergicconjunctivitis, anterior uveitis, cataracts, dry or wet age-relatedmacular degeneration (AMD) or diabetic retinopathy are disclosedcomprising administering a therapeutically effective amount of a surfacetreated microparticle comprising a pharmaceutically active compound to ahost, including a human, in need of such treatment. In one embodiment,the host is a human.

In another embodiment, an effective amount of a surface treatedmicroparticle comprising a pharmaceutically active compound is providedto decrease intraocular pressure (IOP) caused by glaucoma. In analternative embodiment, the surface treated microparticle comprising apharmaceutically active compound can be used to decrease intraocularpressure (IOP), regardless of whether it is associated with glaucoma.

In one embodiment, the disorder is associated with an increase inintraocular pressure (IOP) caused by potential or previously poorpatient compliance to glaucoma treatment. In yet another embodiment, thedisorder is associated with potential or poor neuroprotection throughneuronal nitric oxide synthase (NOS). The surface treated microparticlecomprising a pharmaceutically active compound provided herein may thusdampen or inhibit glaucoma in a host, by administration of an effectiveamount in a suitable manner to a host, typically a human, in needthereof.

Methods for the treatment of a disorder associated with glaucoma,increased intraocular pressure (IOP), optic nerve damage caused byeither high intraocular pressure (IOP) or neuronal nitric oxide synthase(NOS) are provided that includes the administration of an effectiveamount of a surface treated microparticle comprising a pharmaceuticallyactive compound are also disclosed.

In one aspect of the present invention, an effective amount of apharmaceutically active compound as described herein is incorporatedinto a surface treated microparticle, e.g., for convenience of deliveryand/or sustained release delivery. The use of materials in micrometerscale provides one the ability to modify fundamental physical propertiessuch as solubility, diffusivity, and drug release characteristics. Thesemicrometer scale agents may provide more effective and/or moreconvenient routes of administration, lower therapeutic toxicity, extendthe product life cycle, and ultimately reduce healthcare costs. Astherapeutic delivery systems, surface treated microparticles can allowtargeted delivery and sustained release.

In another aspect of the present invention, the surface treatedmicroparticle is coated with a surface agent. The present inventionfurther comprises a method of producing surface treated microparticlescomprising a pharmaceutically active compound. The present inventionfurther comprises methods of using the surface treated microparticlescomprising a pharmaceutically active compound to treat a patient.

In one embodiment, surface treated microparticles including apharmaceutically active compound are obtained by forming an emulsion andusing a bead column as described in, for example, U.S. Pat. No.8,916,196.

In one embodiment, surface treated microparticles including apharmaceutically active compound are obtained by using a vibrating meshor microsieve.

In one embodiment, surface treated microparticles including apharmaceutically active compound are obtained by using slurry sieving.

The processes of producing microspheres described herein are amenable tomethods of manufacture that narrow the size distribution of theresultant particles. In one embodiment, the particles are manufacturedby a method of spraying the material through a nozzle with acousticexcitation (vibrations) to produce uniform droplets. A carrier streamcan also be utilized through the nozzle to allow further control ofdroplet size. Such methods are described in detail in: Berkland, C., K.Kim, et al. (2001). “Fabrication of PLG microspheres with preciselycontrolled and monodisperse size distributions.” J Control Release73(1): 59-74; Berkland, C., M. King, et al. (2002). “Precise control ofPLG microsphere size provides enhanced control of drug release rate.” JControl Release 82(1): 137-147; Berkland, C., E. Pollauf, et al. (2004).“Uniform double-walled polymer microspheres of controllable shellthickness.” J Control Release 96(1): 101-111.

In another embodiment, microparticles of uniform size can bemanufactured by methods that utilize microsieves of the desired size.The microsieves can either be used directly during production toinfluence the size of microparticles formed, or alternatively postproduction to purify the microparticles to a uniform size. Themicrosieves can either be mechanical (inorganic material) or biologicalin nature (organic material such as a membrane). One such method isdescribed in detail in U.S. Pat. No. 8,100,348.

In one embodiment, the surface treated microparticles comprise atherapeutically active compound and have a particle size of 25<Dv50<40μm, Dv90<45 μm.

In one embodiment, the surface treated microparticles comprise atherapeutically active compound and have a particle size of Dv10>10 μm.

In one embodiment, the surface treated microparticles comprise atherapeutically active compound and have only residual solvents that arepharmaceutically acceptable.

In one embodiment, the surface treated microparticles comprise atherapeutically active compound and afford a total release of greaterthan eighty percent by day 14.

In one embodiment, the surface treated microparticles comprise atherapeutically active agent and have syringeability with aregular-walled 26, 27, 28, 29 or 30 gauge needle of 200 mg/ml with noclogging of the syringe.

In one embodiment, the surface treated microparticles comprise atherapeutically active agent and have syringeability with a thin-walled26, 27, 28, 29 or 30 gauge needle of 200 mg/ml with no clogging of thesyringe.

In one embodiment, the surface treated microparticles comprisessunitinib have a particle size of 25<Dv50<40 μm, Dv90<45 μm.

In one embodiment, the surface treated microparticles comprisingsunitinib have a particle size of Dv10>10 μm.

In one embodiment, the surface treated microparticles comprisingsunitinib have only residual solvents that are pharmaceuticallyacceptable.

In one embodiment, the surface treated microparticles comprisingsunitinib afford a total release of greater than eighty percent by day14.

In one embodiment, the surface treated microparticles comprisingsunitinib have syringeability with a regular-walled 26, 27, 28, 29 or 30gauge needle of 200 mg/ml with no clogging of the syringe.

In one embodiment, the surface treated microparticles comprisingsunitinib have syringeability with a thin-walled 26, 27, 28, 29 or 30gauge needle of 200 mg/ml with no clogging of the syringe.

In one embodiment, the surface treated microparticles comprisingsunitinib have an endotoxin level of less than 0.02 EU/mg.

In one embodiment, the surface treated microparticles comprisingsunitinib have a bioburden level of less than 10 CFU/g.

Biodegradable Polymers

The surface treated microparticles can include one or more biodegradablepolymers or copolymers. The polymers should be biocompatible in thatthey can be administered to a patient without an unacceptable adverseeffect. Biodegradable polymers are well known to those in the art andare the subject of extensive literature and patents. The biodegradablepolymer or combination of polymers can be selected to provide the targetcharacteristics of the microparticles, including the appropriate mix ofhydrophobic and hydrophilic qualities, half-life and degradationkinetics in vivo, compatibility with the therapeutic agent to bedelivered, appropriate behavior at the site of injection, etc.

For example, it should be understood by one skilled in the art that bymanufacturing a microparticle from multiple polymers with varied ratiosof hydrophobic, hydrophilic, and biodegradable character that theproperties of the microparticle can be designed for the target use. Asan illustration, a microparticle manufactured with 90 percent PLGA and10 percent PEG is more hydrophilic than a microparticle manufacturedwith 95 percent PLGA and 5 percent PEG. Further, a microparticlemanufactured with a higher content of a less biodegradable polymer willin general degrade more slowly. This flexibility allows microparticlesof the present invention to be tailored to the desired level ofsolubility, rate of release of pharmaceutical agent, and rate ofdegradation.

In certain embodiments, the microparticle includes apoly(α-hydroxyacid). Examples of poly(α-hydroxyacids) include polylactic acid (PLA), polyglycolic acid (PGA),poly(D,L-lactide-co-glycolide)(PLGA), and poly D,L-lactic acid (PDLLA).polyesters, poly (ε-caprolactone), poly (3-hydroxy-butyrate), poly(s-caproic acid), poly (p-dioxanone), poly (propylene fumarate), poly(ortho esters), polyol/diketene acetals, addition polymers,polyanhydrides, poly (sebacic anhydride) (PSA), poly(carboxybis-carboxyphenoxyphosphazene) (PCPP), poly [bis(p-carboxyphenoxy) methane] (PCPM), copolymers of SA, CPP and CPM (asdescribed in Tamat and Langer in Journal of Biomaterials Science PolymerEdition, 3, 315-353, 1992 and by Domb in Chapter 8 of The Handbook ofBiodegradable Polymers, Editors Domb A J and Wiseman R M, HarwoodAcademic Publishers), and poly (amino acids).

In one embodiment, the microparticle includes about at least 90 percenthydrophobic polymer and about not more than 10 percent hydrophilicpolymer. Examples of hydrophobic polymers include polyesters such aspoly lactic acid (PLA), polyglycolic acid (PGA),poly(D,L-lactide-co-glycolide)(PLGA), and poly D,L-lactic acid (PDLLA);polycaprolactone; polyanhydrides, such as polysebacic anhydride,poly(maleic anhydride); and copolymers thereof. Examples of hydrophilicpolymers include poly(alkylene glycols) such as polyethylene glycol(PEG), polyethylene oxide (PEO), and poly(ethylene glycol) amine;polysaccharides; poly(vinyl alcohol) (PVA); polypyrrolidone;polyacrylamide (PAM); polyethylenimine (PEI); poly(acrylic acid);poly(vinylpyrolidone) (PVP); or a copolymer thereof.

In one embodiment, the microparticle includes about at least 85 percenthydrophobic polymer and at most 15 percent hydrophilic polymer.

In one embodiment, the microparticle includes about at least 80 percenthydrophobic polymer and at most 20 percent hydrophilic polymer.

In one embodiment, the microparticle includes PLGA.

In one embodiment, the microparticle includes a copolymer of PLGA andPEG.

In one embodiment, the microparticle includes a copolymer of PLA andPEG.

In one embodiment, the microparticle comprises PLGA and PLGA-PEG, andcombinations thereof.

In one embodiment, the microparticle comprises PLA and PLA-PEG.

In one embodiment, the microparticle includes PVA.

In one embodiment, the microparticles include PLGA, PLGA-PEG, PVA, orcombinations thereof.

In one embodiment, the microparticles include the biocompatible polymersPLA, PLA-PEG, PVA, or combinations thereof.

In one embodiment, the microparticles have a mean size of about 25 μm toabout 30 μm and a median size of about 29 μm to about 31 μm beforesurface treatment.

In one embodiment, the microparticles after surface treatment have aboutthe same mean size and median size. In another embodiment, themicroparticles after surface treatment have a mean size which is largerthan the median size. In another embodiment, the microparticles aftersurface treatment have a mean size which is smaller than the mediansize.

In one embodiment, the microparticles have a mean size of about 25 μm toabout 30 μm or 33 μm and a median size of about 31 μm to about 33 μmafter surface treatment with approximately 0.0075 M NaOH/ethanol to 0.75M NaOH/ethanol (30:70, v:v).

In one embodiment, the microparticles have a mean size of about 25 μm toabout 30 μm or 30 to 33 μm and a median size of about 31 μm to about 33μm after surface treatment with approximately 0.75 M NaOH/ethanol to 2.5M NaOH/ethanol (30:70, v:v).

In one embodiment, the microparticles have a mean size of about 25 μm toabout 30 μm or to 33 μm and a median size of about 31 μm to about 33 μmafter surface treatment with approximately 0.0075 M HCl/ethanol to 0.75M NaOH/ethanol (30:70, v:v).

In one embodiment, the microparticles have a mean size of about 25 μm toabout 30 μm or to 33 μm and a median size of about 31 μm to about 33 μmafter surface treatment with approximately 0.75 M NaOH/ethanol to 2.5 MHCl/ethanol (30:70, v:v).

In one embodiment, a surface-modified solid aggregating microparticle ismanufactured using a wet microparticle.

In one embodiment, the surface-modified solid aggregating microparticlecan release a therapeutic agent over a longer period of time whencompared to a non-surface treated microparticle.

In one embodiment, a surface-modified solid aggregating microparticlecontains less surfactant than a microparticle prior to the surfacemodification.

In one embodiment, a surface-modified solid aggregating microparticle ismore hydrophobic than a microparticle prior to the surface modification.

In one embodiment, a surface-modified solid aggregating microparticle isless inflammatory than a non-surface treated microparticle.

In one embodiment, the agent that removes the surface surfactant of asurface-modified solid aggregating microparticle comprises a solventthat partially dissolves or swells the surface-modified solidaggregating microparticle.

In one aspect of the present invention, an effective amount of apharmaceutically active compound as described herein is incorporatedinto a surface treated microparticle, e.g., for convenience of deliveryand/or sustained release delivery. The use of materials provides theability to modify fundamental physical properties such as solubility,diffusivity, and drug release characteristics. These micrometer scaleagents may provide more effective and/or more convenient routes ofadministration, lower therapeutic toxicity, extend the product lifecycle, and ultimately reduce healthcare costs. As therapeutic deliverysystems, surface treated microparticles can allow targeted delivery andsustained release.

Surfactants

In one embodiment, the manufacture of the microparticle includes asurfactant. Examples of surfactants include, for example,polyoxyethylene glycol, polyoxypropylene glycol, decyl glucoside, laurylglucoside, octyl glucoside, polyoxyethylene glycol octylphenol, TritonX-100, glycerol alkyl ester, glyceryl laurate, cocamide MEA, cocamideDEA, dodecyldimethylamine oxide, and poloxamers. Examples of poloxamersinclude, poloxamers 188, 237, 338 and 407. These poloxamers areavailable under the trade name Pluronic® (available from BASF, MountOlive, N.J.) and correspond to Pluronic® F-68, F-87, F-108 and F-127,respectively. Poloxamer 188 (corresponding to Pluronic® F-68) is a blockcopolymer with an average molecular mass of about 7,000 to about 10,000Da, or about 8,000 to about 9,000 Da, or about 8,400 Da. Poloxamer 237(corresponding to Pluronic® F-87) is a block copolymer with an averagemolecular mass of about 6,000 to about 9,000 Da, or about 6.500 to about8,000 Da, or about 7,700 Da. Poloxamer 338 (corresponding to Pluronic®F-108) is a block copolymer with an average molecular mass of about12,000 to about 18.000 Da, or about 13,000 to about 15,000 Da, or about14,600 Da. Poloxamer 407 (corresponding to Pluronic® F-127) is apolyoxyethylene-polyoxypropylene triblock copolymer in a ratio ofbetween about E101 P56 E101 to about E106 P70 E106, or about E101P56E101, or about E106 P70 E106, with an average molecular mass of about10,000 to about 15,000 Da, or about 12,000 to about 14,000 Da, or about12,000 to about 13,000 Da, or about 12,600 Da.

Additional examples of surfactants that can be used in the inventioninclude, but are not limited to, polyvinyl alcohol (which can behydrolyzed polyvinyl acetate), polyvinyl acetate, Vitamin E-TPGS,poloxamers, cholic acid sodium salt, dioctyl sulfosuccinate sodium,hexadecyltrimethyl ammonium bromide, saponin, TWEEN® 20, TWEEN® 80,sugar esters, Triton X series, L-a-phosphatidylcholine (PC),1,2-dipalmitoylphosphatidycholine (DPPC), oleic acid, sorbitantrioleate, sorbitan mono-oleate, sorbitan monolaurate, polyoxyethylene(20) sorbitan monolaurate, polyoxyethylene (20) sorbitan monooleate,natural lecithin, oleyl polyoxyethylene (2) ether, stearylpolyoxyethylene (2) ether, lauryl polyoxyethylene (4) ether, blockcopolymers of oxyethylene and oxypropylene, synthetic lecithin,diethylene glycol dioleate, tetrahydrofurfuryl oleate, ethyl oleate,isopropyl myristate, glyceryl monooleate, glyceryl monostearate,glyceryl monoricinoleate, cetyl alcohol, stearyl alcohol,cetylpyridinium chloride, benzalkonium chloride, olive oil, glycerylmonolaurate, corn oil, cotton seed oil, sunflower seed oil, lecithin,oleic acid, and sorbitan trioleate.

It should be recognized by one skilled in the art that some surfactantscan be used as polymers in the manufacture of the microparticle. Itshould also be recognized by one skilled in the art that in somemanufacture the microparticle may retain a small amount of surfactantwhich allows further modification of properties as desired.

III. Examples of Disorders to be Treated

In one embodiment, the composition includes a surface treatedmicroparticle which comprises: a surface treated microparticle and apharmaceutically active compound encapsulated in the surface treatedmicroparticle optionally in combination with a pharmaceuticallyacceptable carrier, excipient, or diluent. In one embodiment, thecomposition is a pharmaceutical composition for treating an eye disorderor eye disease.

Non-limiting exemplary eye disorders or diseases treatable with thecomposition include age related macular degeneration, alkaline erosivekeratoconjunctivitis, allergic conjunctivitis, allergic keratitis,anterior uveitis, Behcet's disease, blepharitis, blood-aqueous barrierdisruption, chorioiditis, chronic uveitis, conjunctivitis, contactlens-induced keratoconjunctivitis, corneal abrasion, corneal trauma,corneal ulcer, crystalline retinopathy, cystoid macular edema,dacryocystitis, diabetic keratopathy, diabetic macular edema, diabeticretinopathy, dry eye disease, dry age-related macular degeneration,eosinophilic granuloma, episcleritis, exudative macular edema, Fuchs'Dystrophy, giant cell arteritis, giant papillary conjunctivitis,glaucoma, glaucoma surgery failure, graft rejection, herpes zoster,inflammation after cataract surgery, iridocorneal endothelial syndrome,iritis, keratoconjunctivitis sicca, keratoconjunctivitis inflammatorydisease, keratoconus, lattice dystrophy, map-dot-fingerprint dystrophy,necrotic keratitis, neovascular diseases involving the retina, uvealtract or cornea, for example, neovascular glaucoma, cornealneovascularization, neovascularization resulting following a combinedvitrectomy and lensectomy, neovascularization of the optic nerve, andneovascularization due to penetration of the eye or contusive ocularinjury, neuroparalytic keratitis, non-infectious uveitis ocular herpes,ocular lymphoma, ocular rosacea, ophthalmic infections, ophthalmicpemphigoid, optic neuritis, panuveitis, papillitis, pars planitis,persistent macular edema, phacoanaphylaxis, posterior uveitis,post-operative inflammation, proliferative diabetic retinopathy,proliferative sickle cell retinopathy, proliferative vitreoretinopathy,retinal artery occlusion, retinal detachment, retinal vein occlusion,retinitis pigmentosa, retinopathy of prematurity, rubeosis iritis,scleritis, Stevens-Johnson syndrome, sympathetic ophthalmia, temporalarteritis, thyroid associated ophthalmopathy, uveitis, vernalconjunctivitis, vitamin A insufficiency-induced keratomalacia, vitritis,and wet age-related macular degeneration.

IV. Therapeutically Active Agents to be Delivered

A wide variety of therapeutic agents can be delivered in a long termsustained manner in vivo using the present invention.

A “therapeutically effective amount” of a pharmaceuticalcomposition/combination of this invention means an amount effective,when administered to a patient, to provide a therapeutic benefit such asan amelioration of symptoms of the selected disorder, typically anocular disorder. In certain aspects, the disorder is glaucoma, adisorder mediated by carbonic anhydrase, a disorder or abnormalityrelated to an increase in intraocular pressure (IOP), a disordermediated by nitric oxide synthase (NOS), a disorder requiringneuroprotection such as to regenerate/repair optic nerves, allergicconjunctivitis, anterior uveitis, cataracts, dry or wet age-relatedmacular degeneration (AMD), or diabetic retinopathy.

A “pharmaceutically acceptable salt” is formed when a therapeuticallyactive compound is modified by making an inorganic or organic,non-toxic, acid or base addition salt thereof. Salts can be synthesizedfrom a parent compound that contains a basic or acidic moiety byconventional chemical methods. Generally, such a salt can be prepared byreacting a free acid form of the compound with a stoichiometric amountof the appropriate base (such as Na, Ca, Mg, or K hydroxide, carbonate,bicarbonate, or the like), or by reacting a free base form of thecompound with a stoichiometric amount of the appropriate acid. Suchreactions are typically carried out in water or in an organic solvent,or in a mixture of the two. Generally, non-aqueous media like ether,ethyl acetate, ethanol, isopropanol, or acetonitrile are typical, wherepracticable. Examples of pharmaceutically acceptable salts include, butare not limited to, mineral or organic acid salts of basic residues suchas amines; alkali or organic salts of acidic residues such as carboxylicacids; and the like. The pharmaceutically acceptable salts include theconventional non-toxic salts and the quaternary ammonium salts of theparent compound formed, for example, from non-toxic inorganic or organicacids. For example, conventional non-toxic acid salts include thosederived from inorganic acids such as hydrochloric, hydrobromic,sulfuric, sulfamic, phosphoric, nitric and the like; and the saltsprepared from organic acids such as acetic, propionic, succinic,glycolic, stearic, lactic, malic, tartaric, citric, ascorbic, pamoic,maleic, hydroxymaleic, phenylacetic, glutamic, benzoic, salicylic,mesylic, esylic, besylic, sulfanilic, 2-acetoxybenzoic, fumaric,toluenesulfonic, methanesulfonic, ethane disulfonic, oxalic, isethionic,HOOC—(CH₂)_(n)—COOH where n is 0-4. and the like. Lists of additionalsuitable salts may be found, e.g., in Remington's PharmaceuticalSciences, 17th ed., Mack Publishing Company, Easton, Pa., p. 1418(1985).

In one embodiment, the surface treated microparticle of the presentinvention can comprise a compound for the treatment of glaucoma, forinstance a beta-adrenergic antagonists, a prostaglandin analog, anadrenergic agonist, a carbonic anhydrase inhibitor, aparasympathomimetic agent, a dual anti-VEGF/Anti-PDGF therapeutic or adual leucine zipper kinase (DLK) inhibitor. In another embodiment, thesurface treated microparticle of the present invention can comprise acompound for the treatment of diabetic retinopathy. Such compounds maybe administered in lower doses according to the invention as they may beadministered at the site of the ocular disease.

Examples of beta-adrenergic antagonists include, but are not limited to,timolol (Timoptic®), levobunolol (Betagan®), carteolol (Ocupress®), andmetipranolol (OptiPranolol®).

Examples of prostaglandin analogs include, but are not limited to,latanoprost (Xalatan®), travoprost (Travatan®), bimatoprost (Lumigan®)and tafluprost (Zioptan™).

Examples of adrenergic agonists include, but are not limited to,brimonidine (Alphagan®), epinephrine, dipivefrin (Propine®) andapraclonidine (Lopidine®).

Examples of carbonic anhydrase inhibitors include, but are not limitedto, dorzolamide (Trusopt®), brinzolamide (Azopt®), acetazolamide(Diamox®) and methazolamide (Neptazane®), see structures below:

An example of a parasympathomimetic includes, but is not limited to,pilocarpine.

DLK inhibitors include, but are not limited to, Crizotinib, KW-2449 andTozasertib, see structure below.

Drugs used to treat diabetic retinopathy include, but are not limitedto, ranibizumab (Lucentis®).

In one embodiment, the dual anti-VEGF/Anti-PDGF therapeutic is sunitinibmalate (Sutent®). As de

In one embodiment, the compound is a treatment for glaucoma and can beused as an effective amount to treat a host in need of glaucomatreatment.

In another embodiment, the compound acts through a mechanism other thanthose associated with glaucoma to treat a disorder described herein in ahost, typically a human.

In one embodiment, the therapeutic agent is selected from aphosphoinositide 3-kinase (PI3K) inhibitor, a Bruton's tyrosine kinase(BTK) inhibitor, or a spleen tyrosine kinase (Syk) inhibitor, or acombination thereof.

PI3K inhibitors that may be used in the present invention are wellknown. Examples of PI3 kinase inhibitors include but are not limited toWortmannin, demethoxyviridin, perifosine, idelalisib, Pictilisib,Palomid 529, ZSTK474, PWT33597, CUDC-907, and AEZS-136, duvelisib,GS-9820, BKM 120, GDC-0032 (Taselisib)(2-[4-[2-(2-Isopropyl-5-methyl-1,2,4-triazol-3-yl)-5,6-dihydroimidazo[1,2-d][1,4]benzoxazepin-9-yl]pyrazol-1-yl]-2-methylpropanamide),MLN-1117 ((2R)-1-Phenoxy-2-butanyl hydrogen (S)-methylphosphonate; orMethyl(oxo) ([(2R)-1-phenoxy-2-butanyl]oxy) phosphonium)), BYL-7 19((2S)-N1-[4-Methyl-5-[2-(2,2,2-trifluoro-1,1-dimethylethyl)-4-pyridinyl]-2-thiazolyl]-L2-pyrrolidinedicarboxamide), GSK2126458(2,4-Difluoro-N-{2-(methyloxy)-5-[4-(4-pyridazinyl)-6-quinolinyl]-3-pyridinyl}benzenesulfonamide)(omipalisib), TGX-221((+)-7-Methyl-2-(morpholin-4-yl)-9-(1-phenylaminoethyl)-pyrido[1,2-a]-pyrimidin-4-one),GSK2636771(2-Methyl-1-(2-methyl-3-(trifluoromethyl)benzyl)-6-morpholino-1H-benzo[d]imidazole-4-carboxylicacid dihydrochloride), KIN-193((R)-2-((1-(7-methyl-2-morpholino-4-oxo-4H-pyrido[1,2-a]pyrimidin-9-yl)ethyl)amino)benzoicacid), TGR-1202/RP5264, GS-9820((S)-1-(4-((2-(2-aminopyrimidin-5-yl)-7-methyl-4-mohydroxypropan-1-one),GS-1101(5-fluoro-3-phenyl-2-([S)]-1-[9H-purin-6-ylamino]-propyl)-3H-quinazolin-4-one),AMG-319, GSK-2269557, SAR245409(N-(4-(N-(3-((3,5-dimethoxyphenyl)amino)quinoxalin-2-yl)sulfamoyl)phenyl)-3-methoxy-4methylbenzamide), BAY80-6946(2-amino-N-(7-methoxy-8-(3-morpholinopropoxy)-2,3-dihydroimidazo[1,2-c]quinaz),AS 252424(5-[1-[5-(4-Fluoro-2-hydroxy-phenyl)-furan-2-yl]-meth-(Z)-ylidene]-thiazolidine-2,4-dione),CZ 24832(5-(2-amino-8-fluoro-[1,2,4]triazolo[1,5-a]pyridin-6-yl)-N-tert-butylpyridine-3-sulfonamide),Buparlisib(5-[2,6-Di(4-morpholinyl)-4-pyrimidinyl]-4-(trifluoromethyl)-2-pyridinamine),GDC-0941(2-(H-Indazol-4-yl)-6-[[4-(methylsulfonyl)-1-piperazinyl]methyl]-4-(4-morpholinyl)thieno[3,2-d]pyrimidine),GDC-0980((S)-1-(4-((2-(2-aminopyrimidin-5-yl)-7-methyl-4-morpholinothieno[3,2-d]pyrimidin-6yl)methyl)piperazin-1-yl)-2-hydroxypropan-1-one (also known as RG7422)),SF1126((8S,14S,17S)-14-(carboxymethyl)-8-(3-guanidinopropyl)-17-(hydroxymethyl)-3,6,9,12,15-pentaoxo-1-(4-(4-oxo-8-phenyl-4H-chromen-2-yl)morpholino-4-ium)-2-oxa-7,10,13,16-tetraazaoctadecan-18-oate),PF-05212384(N-[4-[[4-(Dimethylamino)-1-piperidinyl]carbonyl]phenyl]-N-[4-(4,6-di-4-morpholinyl-1,3,5-triazin-2-yl)phenyl]urea)(gedatolisib), LY3023414, BEZ235(2-Methyl-2-{4-[3-methyl-2-oxo-8-(quinolin-3-yl)-2,3-dihydro-1H-imidazo[4,5-c]quinolin-1-yl]phenyl}propanenitrile)(dactolisib), XL-765(N-(3-(N-(3-(3,5-dimethoxyphenylamino)quinoxalin-2-yl)sulfamoyl)phenyl)-3-methoxy-4-methylbenzamide),and GSK1059615(5-[[4-(4-Pyridinyl)-6-quinolinyl]methylene]-2,4-thiazolidenedione),PX886([(3aR,6E,9S,9aR,10R,11aS)-6-[[bis(prop-2-enyl)amino]methylidene]-5-hydroxy-9-(methoxymethyl)-9a,11a-dimethyl-1,4,7-trioxo-2,3,3a,9,10,11-hexahydroindeno[4,5h]isochromen-10-yl]acetate (also known as sonolisib)), LY294002, AZD8186, PF-4989216,pilaralisib, GNE-317, PI-3065, PI-103, NU7441 (KU-57788), HS 173,VS-5584 (SB2343), CZC24832, TG100-115, A66, YM201636, CAY10505, PIK-75,PIK-93, AS-605240, BGT226 (NVP-BGT226), AZD6482, voxtalisib, alpelisib,IC-87114, TGI100713, CH5132799, PKI-402, copanlisib (BAY 80-6946), XL147, PIK-90, PIK-293, PIK-294, 3-MA (3-methyladenine), AS-252424,AS-604850, apitolisib (GDC-0980; RG7422), and the structure described inWO2014/071109 having the formula:

BTK inhibitors for use in the present invention are well known. Examplesof BTK inhibitors include ibrutinib (also known asPCI-32765)(Imbruvica™)(1-[(3R)-3-[4-amino-3-(4-phenoxy-phenyl)pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl]prop-2-en-1-one),dianilinopyrimidine-based inhibitors such as AVL-101 and AVL-291/292(N-(3-((5-fluoro-2-((4-(2-methoxyethoxy)phenyl)amino)pyrimidin-4-yl)amino)phenyl)acrylamide)(Avila Therapeutics) (US Patent publication No 2011/0117073,incorporated herein in its entirety), Dasatinib([N-(2-chloro-6-methylphenyl)-2-(6-(4-(2-hydroxyethyl)piperazin-1-yl)-2-methylpyrimidin-4-ylamino)thiazole-5-carboxamide],LFM-A13 (alpha-cyano-beta-hydroxy-beta-methyl-N-(2,5-ibromophenyl)propenamide), GDC-0834([R-N-(3-(6-(4-(1,4-dimethyl-3-oxopiperazin-2-yl)phenylamino)-4-methyl-5-oxo-4,5-dihydropyrazin-2-yl)-2-methylphenyl)-4,5,6,7-tetrahydrobenzo[b]thiophene-2-carboxamide],CGI-5604-(tert-butyl)-N-(3-(8-(phenylamino)imidazo[1,2-a]pyrazin-6-yl)phenyl)benzamide,CGI-1746(4-(tert-butyl)-N-(2-methyl-3-(4-methyl-6-((4-(morpholine-4-carbonyl)phenyl)amino)-5-oxo-4,5-dihydropyrazin-2-yl)phenyl)benzamide),CNX-774(4-(4-((4-((3-acrylamidophenyl)amino)-5-fluoropyrimidin-2-yl)amino)phenoxy)-N-methylpicolinamide),CTA056(7-benzyl-1-(3-(piperidin-1-yl)propyl)-2-(4-(pyridin-4-yl)phenyl)-1H-imidazo[4,5-g]quinoxalin-6(5H)-one),GDC-0834((R)-N-(3-(6-((4-(1,4-dimethyl-3-oxopiperazin-2-yl)phenyl)amino)-4-methyl-5-oxo-4,5-dihydropyrazin-2-yl)-2-methylphenyl)-4,5,6,7-tetrahydrobenzo[b]thiophene-2-carboxamide),GDC-0837((R)-N-(3-(6-((4-(1,4-dimethyl-3-oxopiperazin-2-yl)phenyl)amino)-4-methyl-5-oxo-4,5-dihydropyrazin-2-yl)-2-methylphenyl)-4,5,6,7-tetrahydrobenzo[b]thiophene-2-carboxamide),HM-71224, ACP-196, ONO-4059 (Ono Pharmaceuticals), PRT062607(4-((3-(2H-1,2,3-triazol-2-yl)phenyl)amino)-2-(((1R,2S)-2-aminocyclohexyl)amino)pyrimidine-5-carboxamidehydrochloride), QL-47(1-(1-acryloylindolin-6-yl)-9-(1-methyl-1H-pyrazol-4-yl)benzo[h][1,6]naphthyridin-2(1H)-one),and RN486(6-cyclopropyl-8-fluoro-2-(2-hydroxymethyl-3-{1-methyl-5-[5-(4-methyl-piperazin-1-yl)-pyridin-2-ylamino]-6-oxo-1,6-dihydro-pyridin-3-yl}-phenyl)-2H-isoquinolin-1-one),and other molecules capable of inhibiting BTK activity, for examplethose BTK inhibitors disclosed in Akinleye et ah, Journal of Hematology& Oncology, 2013, 6:59, the entirety of which is incorporated herein byreference.

Syk inhibitors for use in the present invention are well known, andinclude, for example, Cerdulatinib(4-(cyclopropylamino)-2-((4-(4-(ethylsulfonyl)piperazin-1-yl)phenyl)amino)pyrimidine-5-carboxamide),entospletinib(6-(1H-indazol-6-yl)-N-(4-morpholinophenyl)imidazo[1,2-a]pyrazin-8-amine),fostamatinib([6-({5-Fluoro-2-[(3,4,5-trimethoxyphenyl)amino]-4-pyrimidinyl}amino)-2,2-dimethyl-3-oxo-2,3-dihydro-4H-pyrido[3,2-b][1,4]oxazin-4-yl]methyldihydrogen phosphate), fostamatinib disodium salt (sodium(6-((5-fluoro-2-((3,4,5-trimethoxyphenyl)amino)pyrimidin-4-yl)amino)-2,2-dimethyl-3-oxo-2H-pyrido[3,2-b][1,4]oxazin-4(3H)-yl)methylphosphate), BAY 61-3606(2-(7-(3,4-Dimethoxyphenyl)-imidazo[1,2-c]pyrimidin-5-ylamino)-nicotinamideHCl), RO9021(6-[(1R,2S)-2-Amino-cyclohexylamino]-4-(5,6-dimethyl-pyridin-2-ylamino)-pyridazine-3-carboxylicacid amide), imatinib (Gleevac;4-[(4-methylpiperazin-1-yl)methyl]-N-(4-methyl-3-{[4-(pyridin-3-yl)pyrimidin-2-yl]amino}phenyl)benzamide),staurosporine, GSK143(2-(((3R,4R)-3-aminotetrahydro-2H-pyran-4-yl)amino)-4-(p-tolylamino)pyrimidine-5-carboxamide),PP2(1-(tert-butyl)-3-(4-chlorophenyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine),PRT-060318(2-(((1R,2S)-2-aminocyclohexyl)amino)-4-(m-tolylamino)pyrimidine-5-carboxamide),PRT-062607(4-((3-(2H-1,2,3-triazol-2-yl)phenyl)amino)-2-(((1R,2S)-2-aminocyclohexyl)amino)pyrimidine-5-carboxamidehydrochloride), R112(3,3′-((5-fluoropyrimidine-2,4-diyl)bis(azanediyl))diphenol), R348(3-Ethyl-4-methylpyridine), R406(6-((5-fluoro-2-((3,4,5-trimethoxyphenyl)amino)pyrimidin-4-yl)amino)-2,2-dimethyl-2H-pyrido[3,2-b][,4]oxazin-3(4H)-one),piceatannol (3-Hydroxyresveratol), YM193306 (Singh et al. Discovery andDevelopment of Spleen Tyrosine Kinase (SYK) Inhibitors, J. Med. Chem.2012, 55, 3614-3643), 7-azaindole, piceatannol, ER-27319 (Singh et al.Discovery and Development of Spleen Tyrosine Kinase (SYK) Inhibitors, J.Med. Chem. 2012, 55, 3614-3643 incorporated in its entirety herein),Compound D (Singh et al. Discovery and Development of Spleen TyrosineKinase (SYK) Inhibitors, J. Med. Chem. 2012, 55, 3614-3643 incorporatedin its entirety herein), PRT060318 (Singh et al. Discovery andDevelopment of Spleen Tyrosine Kinase (SYK) Inhibitors, J. Med. Chem.2012, 55, 3614-3643 incorporated in its entirety herein), luteolin(Singh et al. Discovery and Development of Spleen Tyrosine Kinase (SYK)Inhibitors, J. Med. Chem. 2012, 55, 3614-3643 incorporated in itsentirety herein), apigenin (Singh et al. Discovery and Development ofSpleen Tyrosine Kinase (SYK) Inhibitors, J. Med. Chem. 2012, 55,3614-3643 incorporated in its entirety herein), quercetin (Singh et al.Discovery and Development of Spleen Tyrosine Kinase (SYK) Inhibitors, J.Med. Chem. 2012, 55, 3614-3643 incorporated in its entirety herein),fisetin (Singh et al. Discovery and Development of Spleen TyrosineKinase (SYK) Inhibitors, J. Med. Chem. 2012, 55, 3614-3643 incorporatedin its entirety herein), myricetin (Singh et al. Discovery andDevelopment of Spleen Tyrosine Kinase (SYK) Inhibitors, J. Med. Chem.2012, 55, 3614-3643 incorporated in its entirety herein), morin (Singhet al. Discovery and Development of Spleen Tyrosine Kinase (SYK)Inhibitors, J. Med. Chem. 2012, 55, 3614-3643 incorporated in itsentirety herein).

In one embodiment, the therapeutic agent is a MEK inhibitor. MEKinhibitors for use in the present invention are well known, and include,for example, trametinib/GSK1120212(N-(3-{3-Cyclopropyl-5-[(2-fluoro-4-iodophenyl)amino]-6,8-dimethyl-2,4,7-trioxo-3,4,6,7-tetrahydropyrido[4,3-d]pyrimidin-l(2H-yl}phenyl)acetamide),selumetinib(6-(4-bromo-2-chloroanilino)-7-fluoro-N-(2-hydroxyethoxy)-3-methylbenzimidazole-5-carboxamide),pimasertib/AS703026/MSC 1935369((S)-N-(2,3-dihydroxypropyl)-3-((2-fluoro-4-iodophenyl)amino)isonicotinamide),XL-518/GDC-0973(1-({3,4-difluoro-2-[(2-fluoro-4-iodophenyl)amino]phenyl}carbonyl)-3-[(2S)-piperidin-2-yl]azetidin-3-ol),refametinib/BAY869766/RDEAl 19(N-(3,4-difluoro-2-(2-fluoro-4-iodophenylamino)-6-methoxyphenyl)-1-(2,3-dihydroxypropyl)cyclopropane-1-sulfonamide),PD-0325901(N-[(2R)-2,3-Dihydroxypropoxy]-3,4-difluoro-2-[(2-fluoro-4-iodophenyl)amino]-benzamide),TAK733((R)-3-(2,3-Dihydroxypropyl)-6-fluoro-5-(2-fluoro-4-iodophenylamino)-8-methylpyrido[2,3-d]pyrimidine-4,7(3H,8H)-dione),MEK 162/ARRY438162(5-[(4-Bromo-2-fluorophenyl)amino]-4-fluoro-N-(2-hydroxyethoxy)-1-methyl-1H-benzimidazole-6-carboxamide),R05126766 (3-[[3-Fluoro-2-(methylsulfamoylamino)-4-pyridyl]methyl]-4-methyl-7-pyrimidin-2-yloxychromen-2-one),WX-554, R04987655/CH4987655(3,4-difluoro-2-((2-fluoro-4-iodophenyl)amino)-N-(2-hydroxyethoxy)-5-((3-oxo-1,2-oxazinan-2yl)methyl)benzamide),or AZD8330 (2-((2-fluoro-4-iodophenyl)amino)-N-(2hydroxyethoxy)-1,5-dimethyl-6-oxo-1,6-dihydropyridine-3-carboxamide),U0126-EtOH, PD184352 (CI-1040), GDC-0623, BI-847325, cobimetinib,PD98059, BIX 02189, BIX 02188, binimetinib, SL-327, TAK-733, PD318088,and additional MEK inhibitors as described below.

In one embodiment, the therapeutic agent is a Raf inhibitor. Rafinhibitors for use in the present invention are well known, and include,for example, Vemurafinib(N-[3-[[5-(4-Chlorophenyl)-1H-pyrrolo[2,3-b]pyridin-3-yl]carbonyl]-2,4-difluorophenyl]-1-propanesulfonamide),sorafenib tosylate(4-[4-[[4-chloro-3-(trifluoromethyl)phenyl]carbamoylamino]phenoxy]-N-methylpyridine-2-carboxamide;4-methylbenzenesulfonate), AZ628(3-(2-cyanopropan-2-yl)-N-(4-methyl-3-(3-methyl-4-oxo-3,4-dihydroquinazolin-6-ylamino)phenyl)benzamide),NVP-BHG712(4-methyl-3-(1-methyl-6-(pyridin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-ylamino)-N-(3-(trifluoromethyl)phenyl)benzamide),RAF-265(1-methyl-5-[2-[5-(trifluoromethyl)-1H-imidazol-2-yl]pyridin-4-yl]oxy-N-[4-(trifluoromethyl)phenyl]benzimidazol-2-amine),2-Bromoaldisine(2-Bromo-6,7-dihydro-1H,5H-pyrrolo[2,3-c]azepine-4,8-dione), Raf KinaseInhibitor IV(2-chloro-5-(2-phenyl-5-(pyridin-4-yl)-1H-imidazol-4-yl)phenol),Sorafenib N-Oxide(4-[4-[[[[4-Chloro-3(trifluoroMethyl)phenyl]aMino]carbonyl]aMino]phenoxy]-N-Methyl-2pyridinecarboxaMide1-Oxide), PLX-4720, dabrafenib (GSK2118436), GDC-0879, RAF265, AZ 628,SB590885, ZM336372, GW5074, TAK-632, CEP-32496, LY3009120, and GX818(Encorafenib).

In one embodiment, the therapeutic agent is a programmed death protein 1(PD-1) inhibitor, a programmed death protein ligand 1 (PDL1) inhibitor,or a programmed death protein ligand 2 (PDL2) inhibitor. PD-1, PDL1, andPDL2 inhibitors are known in the art, and include, for example,nivolumab (BMS), pembrolizumab (Merck), pidilizumab (CureTech/Teva),AMP-244 (Amplimmune/GSK), BMS-936559 (BMS), and MEDI4736(Roche/Genentech), and MPDL3280A (Genentech).

In one embodiment, a therapeutic agent can be administered in asustained fashion.

In one embodiment, the therapeutic agent is a monoclonal antibody (MAb).Some MAbs stimulate an immune response that destroys cancer cells.Similar to the antibodies produced naturally by B cells, these MAbs“coat” the cancer cell surface, triggering its destruction by the immunesystem. For example, bevacizumab targets vascular endothelial growthfactor(VEGF), a protein secreted by tumor cells and other cells in thetumor's microenvironment that promotes the development of tumor bloodvessels. When bound to bevacizumab, VEGF cannot interact with itscellular receptor, preventing the signaling that leads to the growth ofnew blood vessels. Similarly, cetuximab and panitumumab target theepidermal growth factor receptor (EGFR), and trastuzumab targets thehuman epidermal growth factor receptor 2 (HER-2). MAbs that bind to cellsurface growth factor receptors prevent the targeted receptors fromsending their normal growth-promoting signals. They may also triggerapoptosis and activate the immune system to destroy tumor cells.

Other agents may include, but are not limited to, at least one oftamoxifen, midazolam, letrozole, bortezomib, anastrozole, goserelin, anmTOR inhibitor, a PI3 kinase inhibitor as described above, a dualmTOR-PI3K inhibitor, a MEK inhibitor, a RAS inhibitor, ALK inhibitor, anHSP inhibitor (for example, HSP70 and HSP 90 inhibitor, or a combinationthereof), a BCL-2 inhibitor as described above, apopototic inducingcompounds, an AKT inhibitor, including but not limited to, MK-2206,GSK690693, Perifosine, (KRX-0401), GDC-0068, Triciribine, AZD5363,Honokiol, PF-04691502, and Miltefosine, a PD-1 inhibitor as describedabove including but not limited to, Nivolumab, CT-011, MK-3475,BMS936558, and AMP-514 or a FLT-3 inhibitor, including but not limitedto, P406, Dovitinib, Quizartinib (AC220), Amuvatinib (MP-470),Tandutinib (MLN518), ENMD-2076, and KW-2449, or a combination thereof.Examples of mTOR inhibitors include but are not limited to rapamycin andits analogs, everolimus (Afinitor), temsirolimus, ridaforolimus,sirolimus, and deforolimus. Examples of MEK inhibitors include but arenot limited to tametinib/GSK1120212(N-(3-{3-Cyclopropyl-5-[(2-fluoro-4-iodophenyl)amino]-6,8-dimethyl-2,4,7-trioxo-3,4,6,7-tetrahydropyrido[4,3-d]pyrimidin-l(2H-yl}phenyl)acetamide),selumetinob(6-(4-bromo-2-chloroanilino)-7-fluoro-N-(2-hydroxyethoxy)-3-methylbenzimidazole-5-carboxamide),pimasertib/AS703026/MSC 1935369((S)-N-(2,3-dihydroxypropyl)-3-((2-fluoro-4-iodophenyl)amino)isonicotinamide),XL-518/GDC-0973(1-({3,4-difluoro-2-[(2-fluoro-4-iodophenyl)amino]phenyl}carbonyl)-3-[(2S)-piperidin-2-yl]azetidin-3-ol)(cobimetinib), refametinib/BAY869766/RDEAl19(N-(3,4-difluoro-2-(2-fluoro-4-iodophenylamino)-6-methoxyphenyl)-1-(2,3-dihydroxypropyl)cyclopropane-1-sulfonamide),PD-0325901(N-[(2R)-2,3-Dihydroxypropoxy]-3,4-difluoro-2-[(2-fluoro-4-iodophenyl)amino]-benzamide),TAK733((R)-3-(2,3-Dihydroxypropyl)-6-fluoro-5-(2-fluoro-4-iodophenylamino)-8-methylpyrido[2,3d]pyrimidine-4,7(3H,8H)-dione), MEK 162/ARRY438162(5-[(4-Bromo-2-fluorophenyl)amino]-4-fluoro-N-(2-hydroxyethoxy)-1-methyl-1H-benzimidazole-6carboxamide), R05126766(3-[[3-Fluoro-2-(methylsulfamoylamino)-4-pyridyl]methyl]-4-methyl-7-pyrimidin-2-yloxychromen-2-one),WX-554, R04987655/CH4987655(3,4-difluoro-2-((2-fluoro-4-iodophenyl)amino)-N-(2-hydroxyethoxy)-5-((3-oxo-1,2-oxazinan-2yl)methyl)benzamide), or AZD8330(2-((2-fluoro-4-iodophenyl)amino)-N-(2-hydroxyethoxy)-1,5-dimethyl-6-oxo-1,6-dihydropyridine-3-carboxamide).Examples of RAS inhibitors include but are not limited to Reolysin andsiG12D LODER. Examples of ALK inhibitors include but are not limited toCrizotinib, Ceritinib (Zykadia), AP26113, and LDK378. HSP inhibitorsinclude but are not limited to Geldanamycin or17-N-Allylamino-17-demethoxygeldanamycin (17AAG), and Radicicol.

In certain aspects, the therapeutic agent is an anti-inflammatory agent,a chemotherapeutic agent, a radiotherapeutic, an additional therapeuticagent, or an immunosuppressive agent.

In one embodiment, a chemotherapeutic is selected from, but not limitedto, imatinib mesylate (Gleevac®), dasatinib (Sprycel®), nilotinib(Tasigna®), bosutinib (Bosulif®), trastuzumab (Herceptin®),trastuzumab-DM1, pertuzumab (Perjeta™), lapatinib (Tykerb®), gefitinib(Iressa®), erlotinib (Tarceva®), cetuximab (Erbitux®), panitumumab(Vectibix®), vandetanib (Caprelsa®), vemurafenib (Zelboraf®), vorinostat(Zolinza®), romidepsin (Istodax®), bexarotene (Tagretin®), alitretinoin(Panretin®), tretinoin (Vesanoid®), carfilizomib (Kyprolis™),pralatrexate (Folotyn®), bevacizumab (Avastin®), ziv-aflibercept(Zaltrap®), sorafenib (Nexavar®), sunitinib (Sutent®), pazopanib(Votrient®), regorafenib (Stivarga®), and cabozantinib (Cometriq™).

Additional chemotherapeutic agents include, but are not limited to, aradioactive molecule, a toxin, also referred to as cytotoxin orcytotoxic agent, which includes any agent that is detrimental to theviability of cells, and liposomes or other vesicles containingchemotherapeutic compounds. General anticancer pharmaceutical agentsinclude: vincristine (Oncovin®) or liposomal vincristine (Marqibo®),daunorubicin (daunomycin or Cerubidine®) or doxorubicin (Adriamycin®),cytarabine (cytosine arabinoside, ara-C, or Cytosar®), L-asparaginase(Elspar®) or PEG-L-asparaginase (pegaspargase or Oncaspar®), etoposide(VP-16), teniposide (Vumon®), 6-mercaptopurine (6-MP or Purinethol®),Methotrexate, cyclophosphamide (Cytoxan®), Prednisone, dexamethasone(Decadron), imatinib (Gleevec®), dasatinib (Sprycel®), nilotinib(Tasigna®), bosutinib (Bosulift), and ponatinib (Iclusig™). Examples ofadditional suitable chemotherapeutic agents include but are not limitedto 1-dehydrotestosterone, 5-fluorouracil decarbazine, 6-mercaptopurine,6-thioguanine, actinomycin D, adriamycin, aldesleukin, an alkylatingagent, allopurinol sodium, altretamine, amifostine, anastrozole,anthramycin (AMC)), an anti-mitotic agent, cis-dichlorodiamine platinum(II) (DDP) cisplatin), diamino dichloro platinum, anthracycline, anantibiotic, an antimetabolite, asparaginase, BCG live (intravesical),betamethasone sodium phosphate and betamethasone acetate, bicalutamide,bleomycin sulfate, busulfan, calcium leucouorin, calicheamicin,capecitabine, carboplatin, lomustine (CCNU), carmustine (BSNU),chlorambucil, cisplatin, cladribine, colchicin, conjugated estrogens,cyclophosphamide, cyclothosphamide, cytarabine, cytarabine, cytochalasinB, cytoxan, dacarbazine, dactinomycin, dactinomycin (formerlyactinomycin), daunirubicin HCL, daunorucbicin citrate, denileukindiftitox, Dexrazoxane, Dibromomannitol, dihydroxy anthracin dione,docetaxel, dolasetron mesylate, doxorubicin HCL, dronabinol, E. coliL-asparaginase, emetine, epoetin-α, Erwinia L-asparaginase, esterifiedestrogens, estradiol, estramustine phosphate sodium, ethidium bromide,ethinyl estradiol, etidronate, etoposide citrororum factor, etoposidephosphate, filgrastim, floxuridine, fluconazole, fludarabine phosphate,fluorouracil, flutamide, folinic acid, gemcitabine HCL, glucocorticoids,goserelin acetate, gramicidin D, granisetron HCL, hydroxyurea,idarubicin HCL, ifosfamide, interferon α-2b, irinotecan HCL, letrozole,leucovorin calcium, leuprolide acetate, levamisole HCL, lidocaine,lomustine, maytansinoid, mechlorethamine HCL, medroxyprogesteroneacetate, megestrol acetate, melphalan HCL, mercaptipurine, mesna,methotrexate, methyltestosterone, mithramycin, mitomycin C, mitotane,mitoxantrone, nilutamide, octreotide acetate, ondansetron HCL,paclitaxel, pamidronate disodium, pentostatin, pilocarpine HCL,plimycin, polifeprosan 20 with carmustine implant, porfimer sodium,procaine, procarbazine HCL, propranolol, rituximab, sargramostim,streptozotocin, tamoxifen, taxol, teniposide, tenoposide, testolactone,tetracaine, thioepa chlorambucil, thioguanine, thiotepa, topotecan HCL,toremifene citrate, trastuzumab, tretinoin, valrubicin, vinblastinesulfate, vincristine sulfate, and vinorelbine tartrate.

Additional therapeutic agents can include bevacizumab, sutinib,sorafenib, 2-methoxyestradiol or 2ME2, finasunate, vatalanib,vandetanib, aflibercept, volociximab, etaracizumab (MEDI-522),cilengitide, erlotinib, cetuximab, panitumumab, gefitinib, trastuzumab,dovitinib, figitumumab, atacicept, rituximab, alemtuzumab, aldesleukine,atlizumab, tocilizumab, temsirolimus, everolimus, lucatumumab,dacetuzumab, HLL1, huN901-DM1, atiprimod, natalizumab, bortezomib,carfilzomib, marizomib, tanespimycin, saquinavir mesylate, ritonavir,nelfinavir mesylate, indinavir sulfate, belinostat, panobinostat,mapatumumab, lexatumumab, dulanermin, ABT-737, oblimersen, plitidepsin,talmapimod, P276-00, enzastaurin, tipifarnib, perifosine, imatinib,dasatinib, lenalidomide, thalidomide, simvastatin, celecoxib,bazedoxifene, AZD4547, rilotumumab, oxaliplatin (Eloxatin), PD0332991(palbociclib), ribociclib (LEE01), amebaciclib (LY2835219), HDM201,fulvestrant (Faslodex), exemestane (Aromasin), PIM447, ruxolitinib(INC424), BGJ398, necitumumab, pemetrexed (Alimta), and ramucirumab(IMC-1121B).

In one aspect of the present invention, an immunosuppressive agent isused, preferably selected from the group consisting of a calcineurininhibitor, e.g. a cyclosporin or an ascomycin, e.g. Cyclosporin A(NEORAL®), FK506 (tacrolimus), pimecrolimus, a mTOR inhibitor, e.g.rapamycin or a derivative thereof, e.g. Sirolimus (RAPAMUNE®),Everolimus (Certican®), temsirolimus, zotarolimus, biolimus-7,biolimus-9, a rapalog, e.g. ridaforolimus, azathioprine, campath 1H, aSiP receptor modulator, e.g. fingolimod or an analogue thereof, ananti-IL-8 antibody, mycophenolic acid or a salt thereof, e.g. sodiumsalt, or a prodrug thereof, e.g. Mycophenolate Mofetil (CELLCEPT®), OKT3(ORTHOCLONE OKT3®), Prednisone, ATGAM@, THYMOGLOBULIN®, BrequinarSodium, OKT4, T10B9.A-3A, 33B3.1, 15-deoxyspergualin, tresperimus,Leflunomide ARAVA@, CTLAI-Ig, anti-CD25, anti-IL2R, Basiliximab(SIMULECT®), Daclizumab (ZENAPAX®), mizorbine, methotrexate,dexamethasone, ISAtx-247, SDZ ASM 981 (pimecrolimus, Elidel®), CTLA4lg(Abatacept), belatacept, LFA3lg., etanercept (sold as Enbrel® byImmunex), adalimumab (Humira®), infliximab (Remicade®), an anti-LFA-1antibody, natalizumab (Antegren®), Enlimomab, gavilimomab, antithymocyteimmunoglobulin, siplizumab, Alefacept efalizumab, pentasa, mesalazine,asacol, codeine phosphate, benorylate, fenbufen, naprosyn, diclofenac,etodolac and indomethacin, aspirin and ibuprofen.

Examples of types of therapeutic agents that can be includeanti-inflammatory drugs, antimicrobial agents, anti-angiogenesis agents,immunosuppressants, antibodies, steroids, ocular antihypertensive drugsand combinations thereof. Examples of therapeutic agents includeamikacin, anecortane acetate, anthracenedione, anthracycline, an azole,amphotericin B, bevacizumab, camptothecin, cefuroxime, chloramphenicol,chlorhexidine, chlorhexidine digluconate, clortrimazole, a clotrimazolecephalosporin, corticosteroids, dexamethasone, desamethazone, econazole,eftazidime, epipodophyllotoxin, fluconazole, flucytosine,fluoropyrimidines, fluoroquinolines, gatifloxacin, glycopeptides,imidazoles, itraconazole, ivermectin, ketoconazole, levofloxacin,macrolides, miconazole, miconazole nitrate, moxifloxacin, natamycin,neomycin, nystatin, ofloxacin, polyhexamethylene biguanide,prednisolone, prednisolone acetate, pegaptanib, platinum analogues,polymicin B, propamidine isethionate, pyrimidine nucleoside,ranibizumab, squalamine lactate, sulfonamides, triamcinolone,triamcinolone acetonide, triazoles, vancomycin, anti-vascularendothelial growth factor (VEGF) agents, VEGF antibodies, VEGF antibodyfragments, vinca alkaloid, timolol, betaxolol, travoprost, latanoprost,bimatoprost, brimonidine, dorzolamide, acetazolamide, pilocarpine,ciprofloxacin, azithromycin, gentamycin, tobramycin, cefazolin,voriconazole, gancyclovir, cidofovir, foscarnet, diclofenac, nepafenac,ketorolac, ibuprofen, indomethacin, fluoromethalone, rimexolone,anecortave, cyclosporine, methotrexate, tacrolimus and combinationsthereof.

Examples of immunosuppressive agents are calcineurin inhibitor, e.g., acyclosporin or an ascomycin, e.g., Cyclosporin A (NEORAL®), FK506(tacrolimus), pimecrolimus, a mTOR inhibitor, e.g., rapamycin or aderivative thereof, e.g., Sirolimus (RAPAMUNE®), Everolimus (Certican®),temsirolimus, zotarolimus, biolimus-7, biolimus-9, a rapalog, e.g.,ridaforolimus, azathioprine, campath 1H, a SiP receptor modulator, e.g.,fingolimod or an analogue thereof, an anti IL-8 antibody, mycophenolicacid or a salt thereof, e.g., sodium salt, or a prodrug thereof, e.g.,Mycophenolate Mofetil (CELLCEPT®), OKT3 (ORTHOCLONE OKT3®), Prednisone,ATGAM®, THYMOGLOBULIN®, Brequinar Sodium, OKT4, T10B9.A-3A, 33B3.1,15-deoxyspergualin, tresperimus, Leflunomide ARAVA®, CTLAI-Ig,anti-CD25, anti-IL2R, Basiliximab (SIMULECT®), Daclizumab (ZENAPAX®),mizorbine, methotrexate, dexamethasone, ISAtx-247, SDZ ASM 981(pimecrolimus, Elidel®), CTLA4lg (Abatacept), belatacept, LFA3lg,etanercept (sold as Enbrel® by Immunex), adalimumab (Humira®),infliximab (Remicade®), an anti-LFA-1 antibody, natalizumab (Antegren®),Enlimomab, gavilimomab, antithymocyte immunoglobulin, siplizumab,Alefacept efalizumab, pentasa, mesalazine, asacol, codeine phosphate,benorylate, fenbufen, naprosyn, diclofenac, etodolac and indomethacin,aspirin and ibuprofen.

An aspect of the invention is a method for the treatment of a disorder,comprising administering to a host in need thereof surface-modifiedsolid aggregating microparticles comprising an effective amount of atherapeutic agent, wherein the therapeutic agent containingsurface-modified solid aggregating microparticles are injected into thebody and aggregate in vivo to form at least one pellet of at least 500μm that provides sustained drug delivery for at least one month.

V. Pharmaceutically Acceptable Carriers

Any suitable pharmaceutically acceptable carrier, for example,ophthalmically acceptable viscous carrier, may be employed in accordancewith the invention. The carrier is present in an amount effective inproviding the desired viscosity to the drug delivery system.Advantageously, the viscous carrier is present in an amount in a rangeof from about 0.5 wt percent to about 95 wt percent of the drug deliveryparticles. The specific amount of the viscous carrier used depends upona number of factors including, for example and without limitation, thespecific viscous carrier used, the molecular weight of the viscouscarrier used, the viscosity desired for the present drug delivery systembeing produced and/or used and like factors. Examples of useful viscouscarriers include, but are not limited to, hyaluronic acid, sodiumhyaluronate, carbomers, polyacrylic acid, cellulosic derivatives,polycarbophil, polyvinylpyrrolidone, gelatin, dextrin, polysaccharides,polyacrylamide, polyvinyl alcohol (which can be partially hydrolyzedpolyvinyl acetate), polyvinyl acetate, derivatives thereof and mixturesthereof.

The carrier can also be an aqueous carrier. Example of aqueous carriersinclude, but are not limited to, an aqueous solution or suspension, suchas saline, plasma, bone marrow aspirate, buffers, such as Hank'sBuffered Salt Solution (HBSS), HEPES(4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid), Ringers buffer,ProVisc®, diluted ProVisc®, ProVisc® diluted with PBS, Krebs buffer,Dulbecco's PBS, normal PBS; sodium hyaluronate solution (HA, 5 mg/mL inPBS), simulated body fluids, plasma platelet concentrate and tissueculture medium or an aqueous solution or suspension comprising anorganic solvent.

In one embodiment, the carrier is PBS.

In one embodiment, the carrier is HA, 5 mg/mL in PBS.

In one embodiment, the carrier is ProVisc® diluted with water.

In one embodiment, the carrier is ProVisc® dilution in PBS.

In one embodiment, the carrier is ProVisc® 5-fold diluted with water.

In one embodiment, the carrier is ProVisc® 5-fold dilution in PBS.

In one embodiment, the carrier is ProVisc® 10-fold diluted with water.

In one embodiment, the carrier is ProVisc® 10-fold dilution in PBS.

In one embodiment, the carrier is ProVisc® 20-fold dilution with water.

In one embodiment, the carrier is ProVisc® 20-fold dilution in PBS.

In one embodiment, the carrier is HA, 1.25 mg/mL in an isotonic buffersolution with neutral pH.

The carrier may, optionally, contain one or more suspending agent. Thesuspending agent may be selected from carboxy methylcellulose (CMC),mannitol, polysorbate, poly propylene glycol, poly ethylene glycol,gelatin, albumin, alginate, hydroxyl propyl methyl cellulose (HPMC),hydroxyl ethyl methyl cellulose (HEMC), bentonite, tragacanth, dextrin,sesame oil, almond oil, sucrose, acacia gum and xanthan gum andcombinations thereof.

The carrier may, optionally, contain one or more plasticizers. Thus thecarrier may also include a plasticizer. The plasticizer may, forexample, be polyethylene glycol (PEG), polypropylene glycol, poly(lactic acid) or poly (glycolic acid) or a copolymer thereof,polycaprolactone, and low molecule weight oligomers of these polymers,or conventional plasticizers, such as, adipates, phosphates, phthalates,sabacates, azelates and citrates. The carrier can also include otherknown pharmaceutical excipients in order to improve the stability of theagent.

In one embodiment, one or more additional excipients or deliveryenhancing agents may also be included e.g., surfactants and/orhydrogels, in order to further influence release rate.

VI. Sustained Release of Pharmaceutically Active Compound

The rate of release of the pharmaceutically active compound can berelated to the concentration of pharmaceutically active compounddissolved in the surface treated microparticle. In some embodiments, thepolymeric composition of the surface treated microparticle includesnon-therapeutic agents that are selected to provide a desired solubilityof the pharmaceutically active compound. The selection of the polymericcomposition can be made to provide the desired solubility of thepharmaceutically active compound in the surface treated microparticle,for example, a hydrogel may promote solubility of a hydrophilicmaterial. In some embodiments, functional groups can be added to thepolymer to increase the desired solubility of the pharmaceuticallyactive compound in the surface treated microparticle. In someembodiments, additives may be used to control the release kinetics ofthe pharmaceutically active compound, for example, the additives may beused to control the concentration of the pharmaceutically activecompound by increasing or decreasing the solubility of thepharmaceutically active compound in the polymer so as to control therelease kinetics of the pharmaceutically active compound. The solubilitymay be controlled by including appropriate molecules and/or substancesthat increase and/or decrease the solubility of the dissolved form ofthe pharmaceutically active compound in the surface treatedmicroparticle. The solubility of the pharmaceutically active compoundmay be related to the hydrophobic and/or hydrophilic properties of thesurface treated microparticle and the pharmaceutically active compound.Oils and hydrophobic molecules can be added to the polymer(s) toincrease the solubility of a pharmaceutically active compound in thesurface treated microparticle.

Instead of, or in addition to, controlling the rate of migration basedon the concentration of the pharmaceutically active compound dissolvedin the surface treated microparticle, the surface area of the polymericcomposition can be controlled to attain the desired rate of drugmigration out of the surface treated microparticle comprising apharmaceutically active compound. For example, a larger exposed surfacearea will increase the rate of migration of the pharmaceutically activecompound to the surface, and a smaller exposed surface area willdecrease the rate of migration of the pharmaceutically active compoundto the surface. The exposed surface area can be increased in any numberof ways, for example, by any of castellation of the exposed surface, aporous surface having exposed channels connected with the tear or tearfilm, indentation of the exposed surface, or protrusion of the exposedsurface. The exposed surface can be made porous by the addition of saltsthat dissolve and leave a porous cavity once the salt dissolves. In thepresent invention, these trends can be used to decrease the release rateof the active material from the polymeric composition by avoiding thesepaths to quicker release. For example, the surface area can beminimized, or channels can be avoided.

Where more than one type of polymer is used, each surface treatedmicroparticle may have a different solidifying or setting property. Forexample, the surface treated microparticles may be made from similarpolymers but may have different gelling pHs or different meltingtemperatures or glass transition points.

In order for the surface treated microparticles to form a consolidatedaggregate, the temperature around the particles, for example in thehuman or non-human animal where the composition is administered, isapproximately equal to, or greater than, the glass transitiontemperature (T_(g)) of the polymer particles. At such temperatures thepolymer particles will cross-link to one or more other polymer particlesto form a consolidated aggregate. By cross-link it is meant thatadjacent polymer particles become joined together. For example, theparticles may cross-link due to entanglement of the polymer chains atthe surface of one particle with polymer chains at the surface ofanother particle. There may be adhesion, cohesion or fusion betweenadjacent particles.

Typically, the injectable surface treated microparticles which areformed of a polymer or a polymer blend have a glass transitiontemperature (T_(g)) either close to or just above body temperature (suchas from about 30° C. to 45° C., e.g., from about 35° C. to 40° C., forexample, from about 37° C. to 40° C.). Accordingly, at room temperaturethe surface treated microparticles are below their T_(g) and behave asdiscrete particles, but in the body the surface treated microparticlessoften and interact/stick to themselves. Typically, agglomeration beginswithin 20 seconds to about 15 minutes of the raise in temperature fromroom to body temperature.

The surface treated microparticles may be formed from a polymer whichhas a T_(g) from about 35° C. to 40° C., for example from about 37° C.to 40° C., wherein the polymer is a poly(α-hydroxyacid) (such as PLA,PGA, PLGA, or PDLLA or a combination thereof), or a blend thereof withPLGA-PEG. Typically, these particles will agglomerate at bodytemperature. The injectable surface treated microparticles may compriseonly poly(α-hydroxyacid) particles or other particle types may beincluded. The microparticles can be formed from a blend ofpoly(D,L-lactide-co-glycolide)(PLGA), PLGA-PEG and PVA which has a T_(g)at or above body temperature. In one embodiment, at body temperature thesurface treated microparticles will interact to form a consolidatedaggregate. The injectable microparticle may comprise onlyPLGA/PLGA-PEG/PVA surface treated microparticles or other particle typesmay be included.

The composition may comprise a mixture of temperature sensitive surfacetreated microparticles and non-temperature sensitive surface treatedmicroparticles. Non-temperature sensitive surface treated microparticlesare particles with a glass transition temperature which is above thetemperature at which the composition is intended to be used. Typically,in a composition comprising a mixture of temperature sensitive surfacetreated microparticles and non-temperature sensitive particles the ratioof temperature sensitive to non-temperature sensitive surface treatedmicroparticles is about 3:1, or lower, for example, 4:3. The temperaturesensitive surface treated microparticles are advantageously capable ofcrosslinking to each other when the temperature of the composition israised to or above the glass transition of these microparticles. Bycontrolling the ratio of temperature sensitive surface treatedmicroparticles to non-temperature sensitive surface treatedmicroparticles it may be possible to manipulate the porosity of theresulting consolidated aggregate. The surface treated microparticles maybe solid, that is with a solid outer surface, or they may be porous. Theparticles may be irregular or substantially spherical in shape.

The surface treated microparticles can have a size in their longestdimension, or their diameter if they are substantially spherical, ofless than about 100 μm and more than about 1 μm. The surface treatedmicroparticles can have a size in their longest dimension, or theirdiameter, of less than about 100 μm. The surface treated microparticlescan have a size in their longest dimension, or their diameter, ofbetween about 1 μm and about 40 μm, more typically, between about 20 μmand about 40 μm. Polymer particles of the desired size will pass througha sieve or filter with a pore size of about 40 μm.

Formation of the consolidated aggregate from the composition, onceadministered to a human or non-human animal, typically takes from about20 seconds to about 24 hours, for example, between about 1 minute andabout 5 hours, between about 1 minute and about 1 hour, less than about30 minutes, less than about 20 minutes. Typically, the solidificationoccurs in between about 1 minute and about 20 minutes fromadministration.

Typically, the composition comprises from about 20 percent to about 80percent injectable surface treated microparticle material and from about20 percent to about 80 percent carrier, from about 30 percent to about70 percent injectable surface treated microparticle material and fromabout 30 percent to about 70 percent carrier, e.g., the composition maycomprise from about 40 percent to about 60 percent injectable surfacetreated microparticle material and from about 40 percent to about 60percent carrier; the composition may comprise about 50 percentinjectable surface treated microparticle material and about 50 percentcarrier. The aforementioned percentages all refer to percentage byweight.

The surface treated microparticles are loaded, for example, in thesurface treated microparticle or as a coating on the surface treatedmicroparticle, with a pharmaceutically active compound.

The system of the invention can allow for the pharmaceutically activecompound release to be sustained for some time, for example, release canbe sustained for at least about 2 hours, at least about 4 hours, atleast about 6 hours, at least about 10 hours, at least about 12 hours,at least about 24 hours, at least 48 hours, at least a week, more thanone week, at least a month, at least two months, at least three months,at least four months, at least five months, at least six months, or atleast seven months.

In one embodiment, the surface-modified solid aggregating microparticlesthat produce a pellet in vivo release the therapeutic agent without aburst of more than about 1 percent to about 5 percent of total payloadover a 24 hour period.

In one embodiment, the surface-modified solid aggregating microparticlesthat produce a pellet in vivo release the therapeutic agent without aburst of more than about 10 percent of total payload over a 24 hourperiod.

In one embodiment, the surface-modified solid aggregating microparticlesthat produce a pellet in vivo release the therapeutic agent without aburst of more than about 15 percent of total payload over a 24 hourperiod.

In one embodiment, the surface-modified solid aggregating microparticlesthat produce a pellet in vivo release the therapeutic agent without aburst of more than about 20 percent of total payload over a 24 hourperiod.

In one embodiment, the surface-modified solid aggregating microparticlesthat produce a pellet in vivo release the therapeutic agent without aburst of more than about 1 percent to about 5 percent of total payloadover a 12 hour period.

In one embodiment, the surface-modified solid aggregating microparticlesthat produce a pellet in vivo release the therapeutic agent without aburst of more than about 10 percent of total payload over a 12 hourperiod.

In one embodiment, the surface-modified solid aggregating microparticlesthat produce a pellet in vivo release the therapeutic agent without aburst of more than about 15 percent of total payload over a 12 hourperiod.

In one embodiment, the surface-modified solid aggregating microparticlesthat produce a pellet in vivo release the therapeutic agent without aburst of more than about 15 percent of total payload over a 12 hourperiod.

In one embodiment, the pharmaceutically active compound is released inan amount effective to have a desired local or systemic physiological orpharmacologically effect.

In one embodiment, delivery of a pharmaceutically active compound meansthat the pharmaceutically active compound is released from theconsolidated aggregate into the environment around the consolidatedaggregate, for example, the vitreal fluid.

In one embodiment, a surface treated microparticle comprising apharmaceutically active compound of the invention allows a substantiallyzero or first order release rate of the pharmaceutically active compoundfrom the consolidated aggregate once the consolidated aggregate hasformed. A zero order release rate is a constant release of thepharmaceutically active compound over a defined time; such release isdifficult to achieve using known delivery methods.

VII. Manufacture of Surface Treated Microparticles

Microparticle Formation

Microparticles can be formed using any suitable method for the formationof polymer microparticles known in the art. The method employed forparticle formation will depend on a variety of factors, including thecharacteristics of the polymers present in the drug or polymer matrix,as well as the desired particle size and size distribution. The type ofdrug(s) being incorporated in the microparticles may also be a factor assome drugs are unstable in the presence of certain solvents, in certaintemperature ranges, and/or in certain pH ranges.

Particles having an average particle size of between 1 micron and 100microns are useful in the compositions described herein. In typicalembodiments, the particles have an average particle size of between 1micron and 40 microns, more typically between about 10 micron and about40 microns, more typically between about 20 micron and about 40 microns.The particles can have any shape but are generally spherical in shape.

In circumstances where a monodisperse population of particles isdesired, the particles may be formed using a method which produces amonodisperse population of microparticles. Alternatively, methodsproducing polydispersed microparticle distributions can be used, and theparticles can be separated using methods known in the art, such assieving, following particle formation to provide a population ofparticles having the desired average particle size and particle sizedistribution.

Common techniques for preparing microparticles include, but are notlimited to, solvent evaporation, hot melt particle formation, solventremoval, spray drying, phase inversion, coacervation, and lowtemperature casting. Suitable methods of particle formulation arebriefly described below. Pharmaceutically acceptable excipients,including pH modifying agents, disintegrants, preservatives, andantioxidants, can optionally be incorporated into the particles duringparticle formation.

In one embodiment, surface treated microparticles are prepared usingcontinuous chemistry manufacturing processes. In one embodiment, surfacetreated microparticles are prepared using step-wise manufacturingprocesses.

In one embodiment, microparticles containing a therapeutic agent can beprepared as described in PCT/US2015/065894. In one embodiment, themicroparticles are prepared by:

-   -   (i) dissolving or dispersing the therapeutic agent or its salt        in an organic solvent optionally with an alkaline agent,    -   (ii) mixing the solution/dispersion of step (i) with a polymer        solution that has a viscosity of at least about 300 cPs (or        perhaps at least about 350, 400, 500, 600, 700 or 800 or more        cPs);    -   (iii) mixing the therapeutic agent polymer solution/dispersion        of step (ii) with an aqueous non-acidic or alkaline solution        (for example at least approximately a pH of 7, 8, or 9 and        typically not higher than about 10) optionally with a surfactant        or emulsifier, to form a solvent-laden therapeutic agent        encapsulated microparticle, (iv) isolating the microparticles.        In one embodiment, the therapeutic agent is sunitinib.

It has been found that it may be useful to include the alkaline agent inthe organic solvent. However, as described in PCT/US2015/065894, it hasbeen found that adding an acid to the organic solvent can improve drugloading of the microparticle. Examples demonstrate that polyesters suchas PLGA, PEG-PLGA(PLA) and PEG-PLGA/PLGA blend microparticles displaysustained release of the therapeutic agent or its pharmaceuticallyacceptable salt. Polymer microparticles composed of PLGA and PEGcovalently conjugated to PLGA (M_(w) 45 kDa) (PLGA45k-PEG5k) loaded withthe therapeutic agent were prepared using a single emulsion solventevaporation method. Loading improvement was achieved by increasing thealkalinity of the therapeutic agent in solution, up to 16.1% withPEG-PLGA, which could be further increased by adding DMF, compared toonly 1% with no alkaline added. The therapeutic agent loading wasfurther increased by increasing the pH of the aqueous solution as wellas the polymer solution. Still further significant increases intherapeutic agent loading in the microparticles was achieved byincreasing polymer concentration or viscosity. In one embodiment, thetherapeutic agent is sunitinib.

Solvent Evaporation

In this method, the drug (or polymer matrix and drug) is dissolved in avolatile organic solvent, such as methylene chloride, acetone,acetonitrile, 2-butanol, 2-butanone, t-butyl alcohol, benzene,chloroform, cyclohexane, 1,2-dichloroethane, diethyl ether, ethanol,ethyl acetate, heptane, hexane, methanol, methyl tert-butyl ether,pentane, petroleum ether, iso-propanol, n-propanol, tetrahydrofuran, ormixtures thereof. The organic solution containing the drug is thensuspended in an aqueous solution that contains a surface active agentsuch as poly(vinyl alcohol). The resulting emulsion is stirred untilmost of the organic solvent is evaporated, leaving solid microparticles.The resulting microparticles are washed with water and dried overnightin a lyophilizer. Microparticles with different sizes and morphologiescan be obtained by this method.

Microparticles which contain labile polymers, such as certainpolyanhydrides, may degrade during the fabrication process due to thepresence of water. For these polymers, the following two methods, whichare performed in completely anhydrous organic solvents, can be used.

Oil-in-Oil Emulsion Technique

Solvent removal can also be used to prepare particles from drugs thatare hydrolytically unstable. In this method, the drug (or polymer matrixand drug) is dispersed or dissolved in a volatile organic solvent suchas methylene chloride, acetone, acetonitrile, benzene, 2-butanol,2-butanone, t-butyl alcohol, chloroform, cyclohexane,1,2-dichloroethane, diethyl ether, ethanol, ethyl acetate, heptane,hexane, methanol, methyl tert-butyl ether, pentane, petroleum ether,iso-propanol, n-propanol, tetrahydrofuran, or mixtures thereof. Thismixture is then suspended by stirring in an organic oil (such as siliconoil, castor oil, paraffin oil, or mineral oil) to form an emulsion.Solid particles form from the emulsion, which can subsequently beisolated from the supernatant. The external morphology of spheresproduced with this technique is highly dependent on the identity of thedrug.

Oil-in-Water Emulsion Technique

In this method, the drug (or polymer matrix and drug) is dispersed ordissolved in a volatile organic solvent such as methylene chloride,acetone, acetonitrile, benzene, 2-butanol, 2-butanone, 1-butyl alcohol,chloroform, cyclohexane, 1,2-dichloroethane, diethyl ether, ethanol,ethyl acetate, heptane, hexane, methanol, methyl tert-butyl ether,pentane, petroleum ether, iso-propanol, n-propanol, tetrahydrofuran, ormixtures thereof. This mixture is then suspended by stirring in anaqueous solution of surface active agent, such as poly(vinyl alcohol),to form an emulsion. Solid particles form from the emulsion, which cansubsequently be isolated from the supernatant. The external morphologyof spheres produced with this technique is highly dependent on theidentity of the drug.

As described in PCT/US2015/065894, microparticles with a therapeuticagent can be prepared using the oil-in-water emulsion method. In oneexample, sunitinib microparticles were prepared by dissolving 100 mgPEG-PLGA (5K, 45) in 1 mL methylene chloride, and dissolving 20 mgsunitinib malate in 0.5 mL DMSO and triethylamine. The solutions werethen mixed together, homogenized at 5000 rpm, 1 minute into an aqueoussolution containing 1% polyvinyl alcohol (PVA) and stirred for 2 hours.The particles were collected, washed with double distilled water, andfreeze dried. In another example, sunitinib microparticles were alsoprepared according to PCT/US2015/065894 by dissolving 200 mg PLGA (2A,Alkermers) in 3 mL methylene chloride, and 40 mg sunitinib malate in 0.5mL DMSO and triethylamine. The solutions were then mixed together andhomogenized at 5000 rpm, 1 minute in 1% PVA and stirred for 2 hours. Theparticles were collected, washed with double distilled water, and freezedried.

Spray Drying

In this method, the drug (or polymer matrix and drug) is dissolved in anorganic solvent such as methylene chloride, acetone, acetonitrile,2-butanol, 2-butanone, t-butyl alcohol, benzene, chloroform,cyclohexane, 1,2-dichloroethane, diethyl ether, ethanol, ethyl acetate,heptane, hexane, methanol, methyl tert-butyl ether, pentane, petroleumether, iso-propanol, n-propanol, tetrahydrofuran, or mixtures thereof.The solution is pumped through a micronizing nozzle driven by a flow ofcompressed gas, and the resulting aerosol is suspended in a heatedcyclone of air, allowing the solvent to evaporate from themicrodroplets, forming particles. Particles ranging between 0.1-10microns can be obtained using this method.

Phase Inversion

Particles can be formed from drugs using a phase inversion method. Inthis method, the drug (or polymer matrix and drug) is dissolved in asolvent, and the solution is poured into a strong non solvent for thedrug to spontaneously produce, under favorable conditions,microparticles or nanoparticles. The method can be used to producenanoparticles in a wide range of sizes, including, for example, about100 nanometers to about 10 microns, typically possessing a narrowparticle size distribution.

Coacervation

Techniques for particle formation using coacervation are known in theart, for example, in GB-B-929 406; GB-B-929 40 1; and U.S. Pat. Nos.3,266,987, 4,794,000, and 4,460,563. Coacervation involves theseparation of a drug (or polymer matrix and drug) solution into twoimmiscible liquid phases. One phase is a dense coacervate phase, whichcontains a high concentration of the drug, while the second phasecontains a low concentration of the drug. Within the dense coacervatephase, the drug forms nanoscale or microscale droplets, which hardeninto particles. Coacervation may be induced by a temperature change,addition of a non-solvent or addition of a micro-salt (simplecoacervation), or by the addition of another polymer thereby forming aninterpolymer complex (complex coacervation).

Low Temperature Casting

Methods for very low temperature casting of controlled releasemicrospheres are described in U.S. Pat. No. 5,019,400 to Gombotz et al.In this method, the drug (or polymer matrix and sunitinib) is dissolvedin a solvent. The mixture is then atomized into a vessel containing aliquid non-solvent at a temperature below the freezing point of the drugsolution which freezes the drug droplets. As the droplets andnon-solvent for the drug are warmed, the solvent in the droplets thawsand is extracted into the non-solvent, hardening the microspheres.

Scale Up

The processes for producing microparticles described in the Examples areamenable to scale up by methods known in the art. Examples of suchmethods include U.S. Pat. Nos. 4,822,534; 5,271,961; 5,945,126;6,270,802; 6,361,798; 8,708,159; and U.S. publication 2010/0143479. U.S.Pat. No. 4,822,534 describes a method of manufacture to provide solidmicrospheres that involves the use of dispersions. These dispersionscould be produced industrially and allowed for scale up. U.S. Pat. No.5,271,961 disclosed the production of protein microspheres whichinvolved the use of low temperatures, usually less than 45° C. U.S. Pat.No. 5,945,126 describes the method of manufacture to producemicroparticles on full production scale while maintaining sizeuniformity observed in laboratory scale. U.S. Pat. Nos. 6,270,802 and6,361,798 describe the large scale method of manufacture of polymericmicroparticles whilst maintaining a sterile field. U.S. Pat. No.8,708,159 describes the processing of microparticles on scale using ahydrocyclone apparatus. U.S. publication 2010/0143479 describes themethod of manufacture of microparticles on large scale specifically forslow release microparticles.

XSpray has disclosed a device and the use of supercritical fluids toproduce particles of a size below 10 μM (U.S. Pat. No. 8,167,279).Additional patents to XSpray include U.S. Pat. Nos. 8,585,942 and8,585,943. Sun Pharmaceuticals has disclosed a process for themanufacture of microspheres or microcapsules, WO 2006/123359, hereinincorporated by reference. As an example, Process A involves five stepsthat include 1) the preparation of a first dispersed phase comprising atherapeutically active ingredient, a biodegradable polymer and anorganic solvent 2) mixing the first dispersed phase with an aqueousphase to form an emulsion 3) spraying the emulsion into a vesselequipped to remove an organic solvent and 4) passing the resultingmicrospheres or microcapsules through a first and second screen therebycollecting a fractionated size of the microspheres or microcapsules and5) drying the microspheres or microcapsules.

Xu, Q. et al. have disclosed the preparation of monodispersedbiodegradable polymer microparticles using a microfluidic flow-focusingdevice (Xu, Q., et al “Preparation of Monodispersed BiodegradablePolymer Microparticles Using a Microfluidic Flow-Focusing Device forControlled Drug Delivery”, Small, Vol 5(13): 1575-1581, 2009).

Duncanson, W. J. et al. have disclosed the use of microfluidic devicesto generate microspheres (Duncanson, W. J. et al. “MicrofluidicSynthesis of Monodisperse Porous Microspheres with Size-tunable Pores”,Soft Matter, Vol 8, 10636-10640, 2012).

U.S. Pat. No. 8,916,196 to Evonik describes an apparatus and method forthe production of emulsion based microparticles that can be used inconnection with the present invention.

VIII. Process of Preparation of Surface Treated MicroparticlesAbbreviations

-   DCM, CH₂Cl₂ Dichloromethane-   DL Drug loading-   DMSO Dimethyl sulfoxide-   EtOH Ethanol-   HA Sodium hyaluronate-   hr, h Hour-   min Minute-   NaOH Sodium hydroxide-   NSTMP Non-surface treated microparticles-   PBS Dulbecco's phosphate-buffered saline-   PCL Polycaprolactone-   PEG Polyethylene glycol-   PLA Poly(lactic acid)-   PLGA Poly(lactic-co-glycolic acid)-   PVA Polyvinyl alcohol-   Rpm Revolutions per minute-   RT, r.t. Room temperature-   SD Standard deviation-   STMP Surface treated microparticles-   UV Ultraviolet    General Methods

All non-aqueous reactions were performed under an atmosphere of dryargon or nitrogen gas using anhydrous solvents. The structure ofstarting materials, intermediates, and final products was confirmed bystandard analytical techniques, including NMR spectroscopy and massspectrometry.

Materials

Sodium hydroxide (NaOH, catalog #: S318-1, Fisher Chemical), ethanol(EtOH, catalog #: A405-20, Fisher Chemical), Dulbecco'sphosphate-buffered saline (PBS, catalog #: SH3085003, GE HealthcareHyClone™), sodium hyaluronate (HA, catalog #: AC251770010, AcrosOrganics) and Tween 20 (catalog #: BP337-100, Fisher BioReagents) werepurchased from Fisher Scientific. Polyvinyl alcohol (PVA) (88 percenthydrolyzed, MW approximately 25 kD) (catalog #: 02975) was purchasedfrom Polysciences, Inc. Sunitinib malate was purchased from LCLaboratories (catalog #: S-8803). ProVisc® (10 mg/mL, 0.85 mL, catalog#: 21989, Alcon) was purchased from Besse Medical.Poly(lactic-co-glycolic acid) (PLGA) polymer, poly(lactic-acid) (PLA)polymer, and diblock co-polymers of PLGA and polyethylene glycol(PLGA-PEG) were purchased from the Evonik Corporation (RESOMER Select5050 DLG mPEG 5000 (10 wt percent PEG)). A FreeZone 4.5 liter benchtopfreeze dry system was used for lyophilization.

ProVisc® OVD (Ophthalmic Viscosurgical Device) is a sterile,non-pyrogenic, high molecular weight, non-inflammatory highly purifiedfraction of sodium hyaluronate dissolved in physiological sodiumchloride phosphate buffer. It is FDA approved and indicated for use asan ophthalmic surgical aid. Sodium hyaluronate is a derivative ofhyaluronan for clinical use. Hyaluronan, also known as hyaluronic acid,is a naturally occurring glycosaminoglycan found throughout the bodyincluding in the aqueous and vitreous humors of the eye.

Example 1. Preparation of Biodegradable Non-Surface TreatedMicroparticles (NSTMP) Containing PLGA

Polymer microparticles comprising PLGA and diblock copolymer of PLGA andPEG with or without sunitinib malate were prepared using a singleemulsion solvent evaporation method. Briefly, PLGA (560 mg) and PLGA-PEG(5.6 mg) were co-dissolved in dichloromethane (DCM) (4 mL). Sunitinibmalate (90 mg) was dissolved in dimethyl sulfoxide (DMSO) (2 mL). Thepolymer solution and the drug solution were mixed to form a homogeneoussolution (organic phase). For empty NSTMP, DMSO (2 mL) without drug wasused. For drug-loaded NSTMP, the organic phase was added to an aqueous1% PVA solution in PBS (200 mL) and homogenized at 5,000 rpm for 1minute using an L5M-A laboratory mixer (Silverson Machines Inc., EastLongmeadow, Mass.) to obtain an emulsion. For empty NSTMP, 1 percent PVAsolution in water (200 mL) was used.

The emulsion (solvent-laden microparticles) was then hardened bystirring at room temperature for more than 2 hours to allow the DCM toevaporate. The microparticles were collected by sedimentation andcentrifugation, washed three times in water, and filtered through a40-μm sterile Falcon® cell strainer (Corning Inc., Corning, N.Y.). Thenon-surface treated microparticles (NSTMP) were either used directly inthe surface treatment process or dried by lyophilization and stored as adry powder at −20° C. until used.

Example 2. Surface Treatment of Non-Surface Treated Microparticles(NSTMP) Using NaOH(aq)/EtOH

A pre-chilled solution containing 0.25 M NaOH (aq) and ethanol at apredetermined ratio was added to microparticles in a glass vial understirring in an ice bath at approximately 4° C. to form a suspension at100 mg/mL. The suspension was then stirred for a predetermined time(e.g., 3, 6 or 10 minutes) on ice and poured into a pre-chilledfiltration apparatus to remove the NaOH (aq)/EtOH solution. Themicroparticles were further rinsed with pre-chilled water andtransferred to a 50-mL centrifuge tube. The particles were thensuspended in pre-chilled water and kept in a refrigerator for 30 minutesto allow the particles to settle. Following removal of the supernatant,the particles were resuspended and filtered through a 40-μm cellstrainer to remove large aggregates. Subsequently, the particles werewashed twice with water at room temperature and freeze-dried overnight.Detailed formulation information and conditions of NaOH(aq)/EtOH surfacetreatment experiments are listed in Table 1.

TABLE 1 Detailed batch information on NaOH(aq)/EtOH surface treatedmicroparticles Ratio of 0.25M Batch NaOH (aq) Treatment Microparticlesbefore size to EtOH Time STMP surface treatment (mg) (v/v) (min) ID S-1(99% PLGA 7525 200 30/70 3 S-2 4A, 1% PLGA-PEG) 200 6 S-3 DL = 18.0% 20010 S-4 S-5 (90% PLGA 7525 200 50/50 3 S-6 4A, 10% PLGA-PEG) 200 6 S-7 DL= 18.9% 200 30/70 6 S-8 S-9 (99% PLGA 7525 1000 30/70 3 S-10 4A, 1%PLGA-PEG) DL = 18.3% S-11 (99% PLGA 7525 2300 30/70 3 S-12 4A, 1%PLGA-PEG) DL = 11.1% S-13 (99% PLGA 7525 3600 30/70 3 S-14 4A, 1%PLGA-PEG) DL = 11.9% S-15 (99% PLGA 7525 2000 30/70 3 S-16 4A, 1%PLGA-PEG) DL = 2.15% S-17 (99% PLGA 7525 2000 30/70 3 S-18 4A, 1%PLGA-PEG) DL = 2.21% DL = Drug loading.

Example 3. In Vitro Assessment of Particle Aggregability

Surface treated microparticles (STMP) were suspended in phosphatebuffered saline (PBS) at a concentration of 200 mg/mL. Thirty or fiftymicroliters of the suspension were injected into 1.5-2.0 mL of PBS orsodium hyaluronate solution (HA, 5 mg/mL in PBS) pre-warmed at 37° C. ina 2 mL microcentrifuge tube using a 0.5 mL insulin syringe with apermanent 27-gauge needle (Terumo or Easy Touch brand). Themicrocentrifuge tube was then incubated in a water bath at 37° C. for 2hours. The aggregability of the microparticles was assessed by visualobservation and/or imaging under gentle agitation by inverting and/ortapping and flicking the tubes containing the microparticles.Non-surface treated microparticles (NSTMP) were used as a control.

A successful surface treatment process is expected to result in STMPthat maintain good suspendability, syringeability and injectability.Most importantly, after the injection into PBS or sodium hyaluronate andthe 2 hour incubation at 37° C., the STMP are expected to formconsolidated aggregate(s) that do not break into smaller aggregates orfree floating particles under gentle agitation, a key feature thatdifferentiates STMP from NSTMP and STMP with low aggregability.

Example 4. Effect of Temperature During Surface Treatment onMicroparticle Properties

The effect of temperature on surface treatment was studied by comparingparticles treated at room temperature vs. treated at 4° C. The procedurefor surface treatment at room temperature was identical to the proceduredescribed in Example 2 except that it was conducted at room temperatureinstead of at 4° C.

When the surface treatment process was carried out at room temperaturein a mixture of 0.25 M NaOH and EtOH (v/v: 30/70 or 70/30), theparticles aggregated quickly and irreversibly during surface treatment.In contrast, particles treated at 4° C. in a mixture of NaOH/EtOH at thesame volume ratio did not aggregate during the surface treatment processand maintained good suspendability and injectability uponreconstitution. For surface treatment at room temperature in 0.25 M NaOHwithout EtOH, the particles did not aggregate during the 1 hour surfacetreatment. In addition, STMP treated in NaOH failed to aggregatefollowing incubation at 37° C. In contrast, STMP treated around 4° C.did not aggregate during surface treatment, but aggregated followingincubation at 37° C. After lyophilization and reconstitution in aparticle diluent, the STMP were easily loaded into syringes through a 27gauge needle and injected without needle blockage.

Example 5. Effect of PEG Content on the Aggregability of Surface TreatedMicroparticles

TABLE 2 NSTMP and STMP containing different percentages of PLGA:PLGA-PEGFormulation PLGA PLGA-PEG Surface Treatment # (wt %) (wt %) ConditionS-1 99%  1% None S-3 99%  1% 0.25M NaOH/EtOH (30/70, v/v), 6 min S-5 90%10% None S-8 90% 10% 0.25M NaOH/EtOH (30/70, v/v), 6 min

Two batches of NSTMP (S-1 and S-5) and two batches of STMP (S-3 and S-8)containing different weight percentages of PLGA/PLGA-PEG were surfacetreated following the procedure described below and their aggregabilityin both PBS and HA gel were evaluated.

As listed in Table 2 above, formulation S-3 contained 1% PLGA-PEG andS-8 contained 10% of PLGA-PEG. Samples S-3 and S-8 were individuallytreated in a mixture of 0.25M NaOH and EtOH at a volume ratio of 30/70at 4° C. for 6 minutes. Following injection in PBS and incubation at 37°C. for 2 hours, the microcentrifuge tubes were inverted and theaggregability of the particles was assessed by visual inspection. Asillustrated in FIG. 1 , the NSTMP S-1 and S-started to disperseimmediately after the tubes were inverted, while the STMP, S-3 and S-8,remained aggregated at the bottom of the tubes without dispersionthroughout the entire period of observation (about 10 minutes).

A similar second experiment was conducted by injecting the same particlesuspensions into HA solutions and incubating the samples at 37° C. for 2hours. Immediately after the tubes were inverted, none of the particlesbecame dispersed, including NSTMP; refer to FIG. 2 . This is likely dueto the higher viscosity of HA that prevents particles from diffusingrapidly in the gel solution. Different from S-1 which remainedaggregated throughout the experiment, S-5 started to become dispersed inHA 2 minutes after the tube was inverted. Without wishing to be bound toany one theory, this may be related to the higher PEG content in S-1that affects the interaction between particles and between the particlesurfaces and HA, and thus the diffusion of S-5 in HA was less hinderedthan that of S-1. Though S-8 remained aggregated after injection andincubation in PBS, it appeared more dispersive in HA solution. Incontrast, S-3, which contains less PEG than S-8, was able to aggregatein both PBS and HA solution. These data indicate that the aggregationand dispersion of STMP can be affected by both the particle compositionand properties of the medium where the STMP are injected.

In a third experiment, samples containing S-1, S-2, S-3, S-4, S-5, S-6,S-7 and S-8 were incubated in PBS at 37° C. for 2 hours. After assessingthe aggregability by inverting the tubes, stronger agitation was appliedby tapping the tubes on the bench, which caused the particle aggregatesto detach from the bottom of the tubes. The integrity of the aggregateswas then examined and compared among different formulations. As shown inFIG. 3 , S-3 (1 percent PLGA-PEG) remained as an integrated singleaggregate after detachment from the bottom of the tube. In comparison,though most particles in S-8 (10% PLGA-PEG) remained as one largeaggregate, many dispersed small aggregates or particles were visible inthe tube. The assay with stronger agitation allowed furtherdifferentiation of the aggregability of different particle formulations.Overall, the data suggest that STMP with lower PEG content generallyform stronger and more consolidated aggregates than STMP with higher PEGcontent.

Example 6. Effect of Surface Treatment with PBS/EtOH on Microparticles

Since NaOH is a strong base that may cause partial degradation ofpolymers and lead to rapid modification of the surface properties ofparticles, a neutral phosphate buffered saline (PBS) solution at pH 7.4was evaluated as an alternative to NaOH and the effect of surfacetreatment using PBS/EtOH on microparticles was studied. The surfacetreatment procedure was identical to that described in Example 2, exceptthat the NaOH solution was replaced with PBS (pH 7.4). The experimentwas performed in an ice bath at approximately 4° C. Detailed formulationcomposition and surface treatment conditions are listed in Table 3. Theaggregability of the surface treated microparticles (STMP) was testedfollowing the procedure described in Example 3.

TABLE 3 Formulation composition and conditions of surface treatment withPBS/EtOH Particle ID Batch PBS/ Treatment before Drug size EtOH Timetreatment Composition Loading (mg) (v/v) (min) STMP ID S-11 99% PLGA11.1% 200 30/70 3 S-21 S-19 7525 4A, 1% 11.8% 500 S-22 PLGA-PEG 500 S-23500 6 S-24 S-20   0% 200 6 S-25 200 12 S-26

The results of the aggregability test demonstrated that similar tosurface treatment with NaOH/EtOH, all of the STMP treated with PBS/EtOHwere able to form an aggregate after injection into PBS and incubationfor 2 hours at 37° C. The aggregates appeared stable and resistant togentle agitation; refer to FIG. 4 , a photo of S-21. There was noapparent difference in particle aggregability under in vitro aggregationassay (procedure was conducted as described in Example 3) between theseSTMP and the STMP generated by treatment in NaOH/EtOH. Both drug-loadedSTMP and empty STMP were able to aggregate in PBS, suggesting thesurface treatment process likely has good compatibility with variousparticle formulations with or without drug.

Example 7. Modification of the Surface Treatment Conditions UsingNaOH(Aq)/EtOH

To further optimize the surface treatment conditions with NaOH(aq)/EtOH,the impact of various parameters, such as NaOH concentration,aqueous/EtOH ratio, and treatment time, on surface treatment werestudied (Table 4). It is worth noting that in this Example, the overallmolar concentration of NaOH in the entire aqueous/EtOH mixture was usedas a variable independent of the ratio of aqueous solution to EtOHinstead of using the molarity of NaOH in the aqueous phase only as inExample 2. For example, 0.25M NaOH(aq)/EtOH (v/v: 30/70) in Example 2 isequivalent to 0.075M of NaOH in an aqueous/EtOH (v/v: 30/70) mixture.Thus the volume ratio of aqueous to EtOH was modified from 30/70 to50/50 and 70/30 with the same total amount of NaOH in the mixture. Inaddition, the amount of NaOH was decreased by 10- or 100-fold withoutchanging the ratio of aqueous solution to EtOH. The different treatmenttime was chosen to achieve comparable effectiveness of surfacetreatment. The procedure for surface treatment on microparticles was thesame as Example 2.

TABLE 4 Detailed batch information on modified NaOH(aq)/EtOH STMP NaOHconcentration H₂O/ Treat- Microparticles Batch in H₂O/EtOH EtOH mentbefore surface size mixture ratio Time STMP treatment (mg) (M) (v/v)(min) ID S-27 (99% PLGA 200 0.075 50/50 10 S-28 7525 4A, 1% 200 50/50 20S-29 PLGA-PEG) 200 70/30 15 S-30 DL = 11.3% 200 70/30 30 S-31 200 0.007530/70 3 S-32 200 30/70 10 S-33 200 0.00075 30/70 3 S-34 200 30/70 10S-35

Example 8. Effect of Surface Treatment Using HCl/EtOH on Microparticles

As surface treatment using an aqueous solution of basic pH (Example 2and Example 7) or neutral pH (Example 6) had been tested previously, theeffect of aqueous solution of acidic pH was evaluated in Example 8. HClwas selected as a representative acid. As shown in Table 5,microparticles were treated for 3 minutes in 0.075 M or 0.0075 M of HClin H₂O/EtOH (v/v: 30/70) mixture, respectively. The procedure forHCl/EtOH surface treatment was the same as in Example 2 except that HCl(aq) was used to replace NaOH (aq).

TABLE 5 Detailed batch information of HCl/EtOH treated STMP HClconcentration H₂O/ Treat- Final Microparticles Batch in H₂O/EtOH EtOHment surface before surface size mixture ratio Time treated treatment(mg) (M) (v/v) (min) particles S-27 (99% PLGA 200 0.075 30/70 3 S-367525 4A, 1% 200 0.0075 30/70 3 S-37 PLGA-PEG) DL = 11.3%

Example 9. Surface Treatment on Wet Microparticles

In addition to conducting surface treatment on NSTMP by firstre-suspending NSTMP dry powder in an aqueous solution as illustrated inthe previous examples, the feasibility of surface treatment on NSTMPprior to drying (i.e., “wet” microparticles) was also evaluated. It isexpected to be easier to integrate a surface treatment step using “wet”NSTMP into the entire process of scale-up production of STMP than a stepusing dry powder of NSTMP. After obtaining “wet” NSTMP prior tolyophilization as shown in Example 1, an aliquot of the suspension waslyophilized to determine the particle mass per volume. The particlesuspension was then concentrated or diluted accordingly to reach desiredconcentration and cooled down to desired temperature. Other reagentsneeded for surface treatment were then added to the suspension to reachdesired conditions (e.g., concentration of each chemical reagent) asdescribed in Table 6 to start the surface treatment process. The rest ofthe surface treatment process is the same as described on dry particlesin Example 2. The detailed batch information and experimental conditionsare listed in Table 6.

TABLE 6 Detailed batch information and experimental conditions ofsurface treatment on “wet” microparticles Final surface treatmentsolvent Solute Solute Microparticles Batch (base, concentrationTreatment before surface size acid or in H₂O/EtOH H₂O/EtOH Time STMPtreatment (mg) salt) mixture (M) ratio (v/v) (min) ID S-38 (99% PLGA 450NaOH 0.075 30/70 3 S-39 7525 4A, 1% 450 0.0075 30/70 10 S-40 PLGA-PEG)450 0.075 70/30 15 S-41 DL = 11.6% 450 0.00075 70/30 30 S-42 450 HCl0.0075 30/70 3 S-43 450 KCl 0.075 30/70 20 S-44 450 0.35 30/70 20 S-45

Example 10. Optimized Method for Assessing Particle Aggregability InVitro

To improve the method for assessing particle aggregability in vitro, anorbital shaker was used to replace the manual agitation used in Example3.

Fifty microliters of STMP suspension in PBS at 200 mg/mL was injected in2 mL of PBS pre-warmed at 37° C. in a 16-mm round-bottom glass test tubeusing a 1 mL insulin syringe with a permanent 27-gauge needle (Terumo orEasy Touch brand). The test tube was then incubated in a water bath at37° C. for 2 hours. The aggregability of the microparticles was assessedby visual inspection and/or imaging after shaking for 30 seconds at 400rpm on an orbital shaker (Thermo Scientific™ Multi-Platform Shakers:Catalog No. 13-687-700). The test tube containing particles/aggregateswas then turned horizontally for visual assessment of the particleaggregability. NSTMP were used as a control.

As shown in FIG. 17 , all the STMP in Examples 7 and 8 formed anaggregate after the 2-hour incubation and the aggregates remained mostlyintact following 30-second shaking on an orbital shaker. In contrast,NSTMP in S-27 became fully dispersed following the same agitation. S-12described in Example 2 was also included in this assessment to comparethe aggregability of microparticles treated under different conditions.The results suggest all the modified surface treatment conditions inExamples 7 and 8 resulted in STMP with aggregability similar to that ofS-12.

As shown in FIG. 18 , all the STMP (S-39, S-40, S-41, S-42, S-43, S-44,S-45) in Example 9 formed an aggregate after the 2-hour incubation andthe aggregates remained mostly intact following 30-second shaking on anorbital shaker, while NSTMP (S-38) became fully dispersed following thesame agitation. S-42, S-44 and S-45 appeared to aggregate better thanother STMP samples in FIG. 18 and as well as surface treatment on dryparticle in FIG. 17 . The results demonstrate the success andfeasibility of surface treatment on wet microparticles.

Example 11. Determination of Drug Loading

Drug loading was determined by UV-Vis spectrophotometry. Microparticlescontaining sunitinib (10 mg total weight) were dissolved in anhydrousDMSO (1 mL) and further diluted until the concentration of drug was inthe linear range of the standard curve of UV absorbance of the drug. Theconcentration of the drug was determined by comparing the UV absorbanceto a standard curve. Drug loading is defined as the weight ratio of drugto microparticles.

Example 12. In Vitro Drug Release Study

Microparticles containing sunitinib (10 mg total weight) were suspendedin PBS (4 mL) containing 1% Tween 20 in a 6-mL glass vial and incubatedat 37° C. under shaking at 150 rpm. At predetermined time points, 3 mLof the supernatant was withdrawn after particles settled to the bottomof the vial and replaced with 3 mL of fresh release medium. The drugcontent in the supernatant was determined by UV-Vis spectrophotometry orHPLC. Alternatively, the above procedure can be run at 50° C. todetermine an accelerated in vitro drug release rate as shown in FIG. 5 .

Example 13. Studies on the Effects of Surface Treatment onMicroparticles

Besides aggregability, the effect of surface treatment on otherproperties of microparticles was also studied to fully evaluate thefeasibility of surface treatment. As shown in Table 7, in general, theyield and drug loading of STMP (in Example 2) treated for longer periodsof time were slightly lower than those treated for shorter period oftime, suggesting that at 0.25M NaOH/EtOH (v/v: 3:7), the time window forproducing STMP with high yield and loading is narrow (on the order ofminutes). However, under the modified conditions presented in Example 7,the treatment time can be further extended to tens of minutes withoutreducing DL and yield (Table 7) as well as aggregability (Example 10).STMP treated with HCl(aq)/EtOH in Example 8 maintained the DL prior tosurface treatment with relatively high yield (S-36 and S-37). Inaddition, STMP (S-42, S-44 and S-45) produced by surface treatment onwet microparticles in Example 9 also maintained the DL prior to surfacetreatment with comparable yield as STMP produced by surface treatment ondry particles in Example 7 and 8.

TABLE 7 Yield and drug loading of STMP Drug loading (DL) prior Drugloading after Sample Yield to surface treatment surface treatment S-251% 18.0% 14.2% S-3 50% 18.0% 15.3% S-4 36% 18.0% 6.3% S-6 30% 18.9%15.0% S-7 35% 18.9% 14.7% S-8 28% 18.9% 11.6% S-10 67% 18.3% 18.6% S-1268% 11.1% 11.6% S-14 70% 11.9% 12.0% S-16 56% 2.15% 2.11% S-28 43% 11.3%11.8% S-29 49% 11.3% 11.0% S-30 60% 11.3% 10.1% S-31 61% 11.3% 10.6%S-32 44% 11.3% 12.0% S-33 48% 11.3% 11.5% S-34 49% 11.3% 11.5% S-35 58%11.3% 12.0% S-36 61% 11.3% 10.3% S-37 69% 11.3% 11.6% S-42 44% 11.6%11.2% S-44 50% 11.6% 12.0% S-45 43% 11.6% 12.1%

FIG. 6 illustrates representative in vitro drug release profiles ofNSTMP (S-1) and the corresponding STMP (S-2 and S-3) generated from thesame batch of NSTMP. Overall, the release profiles are similar formicroparticles before and after surface treatment except that theinitial release rate of STMP was lower than that of NSTMP. This suggeststhat under the surface treatment conditions drug molecules that arebound to or near the microparticle surface may have been removed duringthe surface treatment process.

Example 14. Wettability of Surface Treated Microparticles

The wettability of representative batches of STMP and NSTMP wascharacterized using the Washburn method. Briefly, two glass capillarytubes with filter bases were separately filled with equivalent masses ofSTMP and NSTMP dry powder. The bottom of the capillary tubes were theninserted into a beaker with water and water was drawn into the tubesover time due to capillary action. The increase in mass of the tube andthe height of water in the tubes were determined as a function of time.The rate of water absorption was relatively rapid in the tube containingNSTMP, but relatively slow for STMP. Similarly, at the end of the test,the mass increase of the tubes was much higher for NSTMP than for STMP,indicating that the surface modification leads to reduction ofwettability of the microparticles likely due to removal of surfactant orboth surfactant and polymer from particle surface.

Example 15. Preparation of Samples S-10, S-12, S-14, S-16, and S-18 andthe Study of their Drug Release Profiles

Samples S-10 to S-16 and S-18 were prepared at a larger scale of 1 to3.6 grams. The yield and drug loading of these batches are shown inTable 6 above. It is worth noting that the drug loading was notsignificantly changed by surface treatment. The average particle size ofthese STMP samples was similar to that of the corresponding NSTMP priorto surface treatment (data not shown). As shown in FIG. 7 , the releaseprofiles of the STMP prepared at a larger scale (S-14 and S-16) weresimilar to the corresponding NSTMP as well, indicating that the surfacetreatment process had minimal effect on the overall drug release.

Example 16. Injectability and Dosing Consistency of Surface TreatedMicroparticles (STMP)

A suspension of STMP (ST-1-5, approximately 10 percent drug loading) atapproximately 200 mg/mL was prepared by suspending the microparticles in5-fold diluted ProVisc® solution containing 2 mg/mL of HA. After anincubation period of 2 hours at room temperature, 10 μL of the STMPsuspension was loaded into a 50 μL Hamilton syringe with an attached27-gauge needle. Following brief vortexing to fully suspend the STMP,the syringe was held horizontally for 2 minutes and vertically for 2minutes prior to injection into a microcentrifuge tube. The injectionwas repeated using 3 different syringes and each syringe was tested 3times. The STMP in each tube was then dissolved in DMSO and the dose ofdrug was determined by UV-Vis spectrophotometry. As shown in Table 8,excellent dosing consistency between injections using the same syringeand between different syringes was observed, suggesting that the STMPsuspension in diluted ProVisc® remained stable at room temperature for asufficient amount of time to allow consistent dosing of the relativelysmall volume of injection (e.g., 10 μL).

TABLE 8 Injectability and dosing consistency of STMP Average doseStandard Standard Standard Standard Sample UV Dose per syringe deviationdeviation Average dose deviation deviation Name Reading (mg) n = 3 (mg)(mg) (%) n = 9 (mg) (mg) (%) Syringe 1.019 .1966 .1974 .0140 7.0942 1-aSyringe .953 .1838 1-b Syringe 1.098 .2118 1-c Syringe 1.136 .2191 .2058.0122 5.9332 .2031 .0129 6.3345 2-a Syringe 1.052 .2029 2-b Syringe1.012 .1952 2-c Syringe 1.052 .2029 .2062 .0156 7.5633 3-a Syringe 1.157.2232 3-b Syringe .998 .1925 3-c

Example 17. Impact of Microparticle Concentration and Particle Diluenton the Aggregation of Surface Treated Microparticles (STMP)

To investigate the effect of particle concentration and diluent on theaggregation of STMP, STMP suspensions (50 μL) in 5-fold diluted ProVisc®at 2 different microparticle concentrations (100 mg/mL and 200 mg/mL)were injected into 4 mL of PBS or HA solution and incubated at 37° C.for 2 hours.

As illustrated in the top panel of FIG. 8C and FIG. 3D, the STMP at 200mg/mL in diluted ProVisc® were able to form a consolidated aggregate inboth PBS and HA following a 2 hour incubation at 37° C. Compared to 200mg/mL STMP suspended in PBS, the aggregation of 200 mg/mL STMP indiluted ProVisc® appeared slower, but the aggregate became moreconsolidated over time, suggesting the HA molecules in the particlediluent may hinder the contact between STMP and slow down theaggregation process. On the other hand, due to its viscoelasticproperties, HA may help keep particles localized and allow sufficienttime for STMP to form an aggregate. The particle aggregates formed in HAalso appeared to have a more spherical morphology than those formed inPBS, suggesting that if a viscoelastic solution is used as the particlediluent, an optimal range of diluent concentration needs to beidentified to improve the overall performance of STMP aggregation.

After the 2 hour incubation, the strength of the aggregates was testedby shaking the test tubes at 250 rpm on an orbital shaker. Asillustrated in the bottom panel of FIG. 8C and FIG. 8D, the aggregateswere able to endure the shear stress generated by shaking with no orlimited dispersion of microparticles.

In comparison, even though the STMP of 100 mg/mL appeared to form anaggregate in PBS (top panel, FIG. 8A), the aggregate appeared less densethan that of the 200 mg/mL STMP in PBS (top panel, FIG. 8C) and tendedto disaggregate into individual microparticles under agitation (bottompanel, FIG. 8A). In addition, the STMP of 100 mg/mL was not able to formone consolidated aggregate in HA at the end of the 2 hour incubationperiod (top panel, FIG. 8B) and many STMP became dispersed in HA uponshaking at 250 rpm (bottom panel, FIG. 8B). Similar to HA molecules inparticle diluent, the HA molecules in the test medium may furtherdecrease particle-particle contact and reduce the chance of forming aconsolidated aggregate. The results suggest that the aggregability ofSTMP decreases at lower microparticle concentration, possibly due toincreased average particle-particle distance and decreased chance ofdirect contact between particles. The aggregation may also be furtherhindered by other molecules, such as HA, in the test medium.

In summary, the aggregation of STMP can be affected by particleconcentration, particle diluent and the environment into which theparticles are delivered. Overall the data demonstrate that underappropriate conditions, the STMP have good aggregability in differentparticle diluents and test media.

Example 18. Aggregation of Surface Treated Microparticles (STMP) in CowEyes Ex Vivo

To evaluate the aggregability of STMP following intravitreal injectionex vivo, enucleated cow eyes (J. W. Treuth & Sons, Catonsville, Md.)were utilized. The eyes were kept on ice prior to use. Briefly, 30 μL of200 mg/mL STMP, S-10, suspended in 5-fold diluted ProVisc® was injectedinto the central vitreous of cow eyes using a 0.5 mL insulin syringe(Terumo) with a 27-gauge needle and three injections were performed ineach cow eye at different locations. After a 2 hour incubation at 37°C., the eyes were cut open and the aggregates of STMP were examinedusing a dissecting microscope. As shown in FIG. 9 , the injected STMPformed consolidated aggregates in cow vitreous and no apparent particledispersion was observed.

Example 19. Aggregation of Surface Treated Microparticles (STMP) inRabbit Eyes In Vivo

To study the aggregation of surface treated microparticles in rabbiteyes in vivo, 50 μL of 200 mg/mL STMP, S-10, suspended in PBS (FIG. 10A)or 5-fold diluted ProVisc® (FIG. 10B) were injected to the centralvitreous of Dutch Belted rabbit eyes using a 0.5 mL insulin syringe(Terumo) with a 27 gauge needle. Four days after the dosing, the rabbitswere sacrificed and the eyes were nucleated and frozen immediately. Thefrozen eyes were cut into halves and the posterior half of the eye wasthawed at room temperature for 3 minutes to allow isolation of thevitreous from the eye cup, as shown in the left photo of FIG. 10A andFIG. 10B. The frozen vitreous containing particles was placed in acassette to allow the vitreous to thoroughly thaw. The aggregates ofSTMP in the vitreous could be easily separated from vitreous usingforceps, proving the formation of consolidated STMP aggregates in rabbiteyes.

Example 20. Distribution, Tolerability and Pharmacokinetics ofSunitinib-Encapsulated Surface Treated Microparticles (STMP) Followingan Intravitreal (IVT) Injection in Rabbits

The distribution and tolerability of STMP and NSTMP were studied inpigmented New Zealand rabbits (F1) following an intravitreal injectionof the microparticles. ProVisc® was diluted 5-fold in PBS and used as adiluent to prepare particle suspensions of about 200 mg/mL forinjection. Detailed study groups and conditions are presented in Table9.

Complete ocular examinations were performed for up to 7 months after thedosing, using a slit lamp biomicroscope and an indirect ophthalmoscope,to evaluate ocular surface morphology, anterior segment and posteriorsegment inflammation, cataract formation, and retinal changes. A retinallens was used to examine the location, morphology and distribution ofthe microspheres in vitreous. Histological analysis was also performedon enucleated and fixed eyes for up to 7 months. At pre-determined timepoints for up to 7 months, the drug levels of sunitinib (ng/g) invarious ocular tissues (e.g. vitreous, retina, and RPE/choroid) andplasma were also analyzed. FIG. 1 1A illustrates a representative1-month histology image following injection with surface treatedmicroparticles (STMP) and FIG. 11B illustrates a representative 1-monthhistology images following injection with non-surface treatedmicroparticles (NSTMP).

TABLE 9 Detailed information on rabbit study groups and dosingconditions Microsphere *SM Microsphere Microsphere Type Group # MassDose Drug Loading Injection Volume With Drug- #1 2 mg 0.2 mg 10% 10 uLsurface loaded #2 10 mg 1.0 mg 10% 50 uL treatment #3 10 mg 0.2 mg  2%50 uL Empty #7 2 and 10 mg None None 10 uL (Left eye) 50 uL (Right eye)Without Drug- #4 2 mg 0.2 mg 10% 10 uL surface loaded #5 10 mg 1.0 mg10% 50 uL treatment #6 10 mg 0.2 mg  2% 50 uL Empty #8 2 and 10 mg NoneNone 10 uL (Left eye) 50 uL (Right eye) *SM = Sunitinib Malate Dose

Immediately following dosing, the microspheres remained localized at thesite of injection in the vitreous as a depot for all the injections. At1 and 2 months, fundus examination using a retina lens showed that inthe eyes injected with STMP, most particle injections remainedconsolidated in the vitreous without dispersion and no vision impairmentor disturbance was observed. In contrast, particle dispersion was morecommonly observed in the eyes injected with NSTMP.

Histological analysis for up to 7 months showed that overall theinjections were well tolerated with minimal evidence of ocularinflammation or toxicity. No evidence of retinal toxicity (thinning anddegeneration, etc.) was observed with any treatment. With STMP, the onlyeyes with observed inflammation were those with injection-related lenstrauma/cataract and associated secondary lens-induced uveitis, which isbelieved to be associated with the injection procedure and not the STMP;no other evidence of inflammation in eyes dosed with surface treatedmicrospheres was observed (FIG. 11 , left). In some of the eyes dosedwith NSTMP, very mild, but present, inflammation in the vitreous thatmay be associated with the NSTMP was observed (FIG. 11 , right). Theresults suggest that surface treatment not only reduces the chance ofparticle dispersion in the vitreous that can cause visual impairment ordisturbance, but it may also reduce potential intraocular inflammationassociated with microspheres and improve the overall safety of thetreatment.

As shown in FIGS. 14, 15, and 16 , the sunitinib levels in the retina orRPE/choroid of rabbits receiving STMP containing 1 or 0.2 mg ofsunitinib malate were above the K_(i) for sunitinib against VEGFR andPDGFR at 1, 2, and 4 months, respectively. Low levels of sunitinib weredetected in plasma only at 1 and 2 months.

Example 21. Determination of Drug Purity and Impurities in Particles

Sample S-12 (10.5 mg) was measured into an amber vial.N,N-dimethylacetamide (0.3 mL) and acetonitrile (0.6 mL) were added todissolve the particles. Water (2.1 mL) was added and the mixture wasthoroughly mixed. The final concentration of particles inN,N-dimethylacetamide/acetonitrile/water (v/v 1:2:7) mixture was 3.5mg/mL. The purity of active compound in STMP S-12 was determined by HPLCand is reported in Table 10. The results suggest that the surfacetreatment did not affect the purity of encapsulated drug.

TABLE 10 HPLC analysis of drug purity in STMP Peak Retention Area Numbertime (%) 1 0.24 0.157 2 0.78 0.283 3 0.82 0.044 4 1.00 99.39 5 1.120.046 6 1.41 0.084

Example 22. Measurement of Average Size and Size Distribution of SurfaceTreated Microparticles (STMP)

Several milligrams of S-12 were suspended in water. The mean particlesize and distributions were determined using a Coulter Multisizer IV(Beckman Coulter, Inc., Brea, Calif.). The distribution shown in FIG. 12has the following statistics: D10 of 20.98 μm, D50 of 32.32 μm, D90 of41.50 μm, mean of 31.84 μm, and standard deviation of 8.07 μm.

Example 23. Determination of Endotoxin Level in Particle Suspension

Microparticles (5-10 mg, S-12) were added to a sterile vial in abiosafety cabinet. The particles were suspended in endotoxin-free PBS.Using a ToxinSensor™ chromogenic LAL endotoxin assay kit (GenScript USAInc., Piscataway, N.J.) and the instructions provided by themanufacture, the sample's total level of endotoxin was measured. S-12had a low endotoxin level of less than 10 μEU/mg.

Example 24. Toxicity Studies

An acute, non-GLP IVT study was conducted to evaluate the oculartolerability and toxicity of sunitinib malate (free drug) for up to 7days following a single IVT injection. Sunitinib malate was formulatedin phosphate buffered saline and injected bilaterally (0.1 mL) at 0.125or 1.25 mg per eye. At the 1.25 mg/eye dose, histologically significantfindings related to sunitinib included residual test article, lenticularvacuoles/degeneration, mild to minimal inflammatory cell infiltration invitreous, retinal degeneration, detachment, and necrosis. Notoxicologically significant findings were observed at the 0.125 mg/eyedose, which is considered the no-observed-adverse-effect-level (NOAEL)dose.

FIG. 13A, FIG. 13B, and FIG. 13C illustrate select PK profiles forsunitinib malate in the retina, vitreous, and plasma, respectively, frompigmented rabbits.

Example 25. Preparation of Sunitinib Microparticles (not SurfaceTreated)

PLGA (555 mg) and PLGA-PEG5K (5.6 mg) were dissolved in DCM (4 mL).Sunitinib malate (90 mg) was dissolved in DMSO (2 mL). The polymer anddrug solutions were then mixed. The resulting reaction mixture wasfiltered through a 0.22 μm PTFE syringe filter. The resulting reactionmixture was diluted with 1% PVA in PBS (200 mL) in a 250 mL beaker andthen homogenized at 5,000 rpm for 1 minute. (The polymer/drug solutionwas poured into the aqueous phase using homogenization conditions andhomogenized at 5,000 rpm for 1 minute) The reaction was next stirred at800 rpm at room temperature for 3 hours in a biosafety cabinet. Theparticles were allowed to settle in the beaker for 30 minutes andapproximately 150 mL of the supernatant was decanted off. Themicroparticle suspension underwent centrifugation at 56×g for 4.5minutes, the solvent was removed, and the microparticles were thenwashed three times with water. The microparticle size and sizedistribution was determined using a Coulter Multisizer IV prior tolyophilization. The microparticles were lyophilized using a FreeZone 4.5liter benchtop lyophilizer. Light exposure was avoided throughout theentire process.

Example 26. General Procedure for the Preparation of Surface TreatedSunitinib Microparticles

Microparticle dry powder was weighed and placed in a small beaker and astirring bar was added. The beaker was placed in an ice bath and cooledto about 4° C. A NaOH/EtOH solution was prepared by mixing NaOH in water(0.25M) with EtOH at 3:7 (v/v) and cooling to about 4° C. The coldNaOH/EtOH solution was added with stirring to the beaker containing themicroparticles to afford a particle suspension of 100 mg/mL. Thesuspension was stirred for 3 minutes at about 4° C. and poured into afiltration apparatus to quickly remove the NaOH/EtOH solution. (Thefiltration apparatus needed to be pre-chilled in a −20° C. freezer priorto use.) Following filtration, the microparticles were rinsed in thefiltration apparatus with ice cold deionized water and transferred to 50mL centrifuge tubes. Each 50 mL centrifuge tube with filled with coldwater to afford a 40 mL particle suspension at a concentration of 5-10mg/mL. The centrifuge tubes were placed in a regenerator and theparticles were allowed to settle for 30 minutes. The supernatant wasthen decanted. The particles were resuspended in cold water and filteredthrough a 40 μm cell strainer to remove any large aggregates. Theparticles were collected by centrifugation (56×g for 4.5 minutes) andwashed twice with water. The product was lyophilized using a FreeZone4.5 liter benchtop lyophilizer. The surface treatment process wasconducted at approximately 4° C. and light exposure was avoidedthroughout the entire process.

Example 27. Method for Determining Accelerating In Vitro Drug Release at50° C.

Microparticles (10 mg) were added to glass scintillation vials. Fourmilliliters of the release medium (1% Tween 20 in 1×PBS at pH 7.4) wasadded into the vials and the mixtures were vortexed. The vials wereshaken on an orbital shaker at 150 rpm in a Fisher general-purposeincubator at 50° C. At pre-determined time points, the appropriate vialwas cooled and the particles were allowed to settle for 10 minutes.Release medium (3 mL) was then carefully removed from the top of thevial and replaced with fresh release medium (3 mL). The vial was thenreturned to the orbital shaker and the amount of drug in the releasemedium was measured by UV spectroscopy. The concentration of drug wasdetermined by comparing to a standard curve for the drug.

Example 28. Preparation of Biodegradable Surface-Treated Microparticles(STMP) Comprising PLA

NSTMP were first produced similarly as described in Example 1. Briefly,PLA and PLGA-PEG were co-dissolved in dichloromethane (DCM) andsunitinib malate was dissolved in dimethyl sulfoxide (DMSO). The polymersolution and the drug solution were mixed to form a homogeneous solution(organic phase). For empty microparticles, DMSO without drug was used.The organic phase was added to an aqueous 1% PVA solution andhomogenized at 5,000 rpm for 1 minute using an L5M-A laboratory mixer(Silverson Machines Inc., East Longmeadow, Mass.) to obtain an emulsion.The emulsion (solvent-laden microparticles) was then hardened bystirring at room temperature for more than 2 hours to allow the DCM toevaporate. The microparticles were collected by sedimentation andcentrifugation, washed three times in water, and filtered through a40-μm sterile Falcon® cell strainer (Corning Inc., Corning, N.Y.). Thenon-surface-treated microparticles (NSTMP) were either used directly inthe surface treatment process or dried by lyophilization and stored as adry powder at −20° C. until used.

A pre-chilled solution containing NaOH and ethanol was added tomicroparticles in a glass vial under stirring in an ice bath atapproximately 4° C. to form a suspension. The suspension was thenstirred for a predetermined time on ice and poured into a pre-chilledfiltration apparatus to remove the NaOH (aq)/EtOH solution. Themicroparticles were further rinsed with pre-chilled water andtransferred to a 50-mL centrifuge tube. The STMP were then suspended inpre-chilled water and kept in a refrigerator for 30 minutes to allow theparticles to settle. Following removal of the supernatant, the particleswere resuspended and filtered through a 40-μm cell strainer to removelarge aggregates. Subsequently, the particles were washed twice withwater at room temperature and freeze-dried overnight.

TABLE 11 Detailed formulation information of STMP comprising PLA NSTMPSurface Treatment STMP Aqueous Particle Treatment ID Polymer Drug PhaseMixing Solution Conc. Time S-46 800 mg PLA 100 100 mg 200 mL of 5000 rpm1 min 0.075M 200 mg/mL 3 min 4A and 8 mg sunitinib malate 1% PVA in NaOHand PLGA-PEG in 4 mL in 1 mL DMSO PBS 50% EtOH DCM S-47 800 mg PLA 100 1mL DMSO 200 mL of 5000 rpm 1 min 0.075M 200 mg/mL 3 min 4A and 8 mg 1%PVA in NaOH and PLGA-PEG in 4 mL water 50% EtOH DCM S-48 640 mg PLA 1002 mL DMSO 200 mL of 5000 rpm 1 min 0.075M 200 mg/mL 3 min 4A and 6.4 mg1% PVA in NaOH and PLGA-PEG in 4 mL water 50% EtOH DCM

The in vitro aggregability of the STMP was characterized similarly asdescribed in Example 3. Briefly, STMP were suspended in PBS at 200 mg/mLand 30-50 uL of the suspension was injected into 1.5-2.0 mL of PBSpre-warmed at 37° C. After incubation at 37° C. for 2 hours, theaggregability of the microparticles was assessed by visual observationand/or imaging following gentle mechanical agitation. Overall all STMPdescribed in Table 11 were able to aggregate upon incubation at 37° C.for 2 hours.

Example 29. Distribution, Tolerability and Pharmacokinetics ofSunitinib-Encapsulated STMP Comprising PLA Following an Intravitreal(IVT) Injection in Rabbits

Sunitinib-encapsulated STMP comprising PLA were suspended in ProVisc®diluted 5-fold in PBS to achieve a target dose of 1 mg sunitinib malatein a 50 uL particle suspension. The tolerability and pharmacokineticswere studied in pigmented New Zealand rabbits (F1) following anintravitreal injection of the STMP suspension. At pre-determined timepoints after the dosing, complete ocular examinations were performed andthe drug levels of sunitinib (ng/g) in various ocular tissues (e.g.vitreous, retina, and RPE/choroid) were also analyzed (FIG. 19 ).

Ocular examinations for up to 6 months showed that the STMP were welltolerated in rabbit eyes and remained consolidated in the vitreouswithout dispersion and no vision impairment or disturbance was observed.As shown in FIG. 19 , the sunitinib levels in retina or RPE/choroid ofrabbits receiving STMP containing 1 mg of sunitinib malate were abovethe K, for sunitinib against VEGFR and PDGFR at 10 days and 3 months.

Example 30. Production of Surface-Treated Microparticles (STMP) on aLarger Scale (100 g and Higher)

NSTMP were produced using a continuous flow, oil-in-water emulsificationmethod. The scale of the pilot batches was 100-200 g. A dispersed phase(DP) and a continuous phase (CP) were first prepared. For placebomicroparticles, the DP was prepared by co-dissolving PLGA and PLGA-PEGpolymers in DCM. The CP was a 0.25% PVA solution in water. Fordrug-loaded microparticles, the DP was prepared by dissolving sunitinibmalate in DMSO and mixing with the polymer solution in DCM. The CP was a0.25% PVA solution in PBS (pH approximately 7). Detailed formulationparameters are listed in Table 12. An emulsion was produced by mixingthe DP and the CP using a high shear inline mixer. The solvents in theDP were diluted by the CP, causing the emulsion droplets to solidify andbecome polymer microparticles. The microparticles were then washed withwater using the volume exchange principle with the addition of freshwater and removal of solvent-containing water with a hollow fiberfilter. The washed microparticles were subsequently suspended in asolution containing NaOH and ethanol for surface modification of theNSTMP. This step was performed in a jacketed vessel and the temperatureof the suspension was maintained around 8° C. Several surface treatmentconditions have been tested as shown in Table 12. Following additionalwashing in water and analysis of the microparticle and drugconcentration of in-process samples, the STMP suspension was adjusted totarget concentration prior to filling of glass vials. In some batches,mannitol was added to the final suspension. The vials were thenlyophilized and sealed. The manufacturing process can be completedaseptically and the final product in vials may also be terminallysterilized by E-Beam or gamma irradiation.

TABLE 12 Formulation and process parameters of STMP produced on largerscale NSTMP DP Mixing Surface Treatment PLGA PLGA- Sunitinib DMSO speedTime NaOH 7525 4A (g) PEG5k (g) DCM (g) Malate (g) (g) (rpm) (min) EtOH(mM) Excipient 86 0.86 640 16.5 260 4000 30 30% 0.53 86 0.86 640 16.5260 4000 60 30% 75 86 0.86 640 15.3 260 4000 30 40% 0.075 86 0.86 64015.3 260 4000 30 40% 0.75 86 0.86 640 15.3 260 4000 30 40% 0.75 86 0.86640 15.3 260 4000 30 40% 0.75 86 0.86 640 3600 30 50% 0.75 86 0.86 6403600 30 40% 0.75 86 0.86 640 260 3300 30 50% 0.75 86 0.86 640 15.3 2604000 30 40% 0.75 86 0.86 640 3600 30 60% 0.75 86 0.86 640 15.3 260 400030 40% 0.75 86 0.86 640 3600 30 70% 0.75 86 0.86 640 15.3 260 4000 3050% 0.75 Mannitol 86 0.86 640 15.3 260 4000 30 60% 0.75 Mannitol 86 0.86640 15.3 260 4000 30 70% 0.75 Mannitol 172 1.72 1280 30.6 520 4000 3070% 0.75 Mannitol 172 1.72 1280 3600 30 70% 0.75 172 1.72 1280 3600 2570% 0.75 172 1.72 1280 30.6 520 4000 30 60% 0.75 Mannitol 172 1.72 128030.6 520 4000 30 60% 0.75 Mannitol 172 1.72 1280 3600 25 70% 0.75Mannitol 172 1.72 1280 30.6 520 3800 30 60% 0.75 Mannitol 172 1.72 128030.6 520 4000 30 60% 0.75 172 1.72 1280 30.6 520 4000 30 60% 0.75

The in vitro aggregability of the STMP was characterized by a similarmethod to that in Example 3. Briefly, STMP was suspended in PBS at 200mg/mL and 30-50 uL of the suspension was injected into 1.5-2.0 mL of PBSpre-warmed to 37° C. After incubation at 37° C. for 2 hours, theaggregability of the microparticles was assessed by visual observationand/or imaging following gentle mechanical agitation. In general, allSTMP treated with a solution containing 0.75 mM NaOH and EtOH of 40, orhigher were able to aggregate upon incubation at 37° C. Followingsuspension in hyaluronate solution and injection in PBS, STMP treatedwith a higher concentration of EtOH showed a higher tendency offloatation in PBS, suggesting reduced wettability and increased surfacehydrophobicity as a result of the surface treatment.

This specification has been described with reference to embodiments ofthe invention. However, one of ordinary skill in the art appreciatesthat various modifications and changes can be made without departingfrom the scope of the invention as set forth herein. Accordingly, thespecification is to be regarded in an illustrative rather than arestrictive sense, and all such modifications are intended to beincluded within the scope of invention.

We claim:
 1. Surface-modified aggregating microparticles comprising atleast one biodegradable polymer, a surfactant, and a therapeutic agentthat is not covalently bound to the biodegradable polymer wherein themicroparticles have a mean diameter between 10 μm and 60 μm that: (i)contain from about 0.001 percent to about 1 percent surfactant and havebeen surface-modified to contain less surfactant than a microparticleprior to the surface modification wherein the surface has been modifiedat a temperature less than about 18° C.; and (ii) aggregate in vivo toform at least one pellet of at least 500 μm in vivo capable of sustaineddrug delivery in vivo for at least one month.
 2. The surface modifiedsolid aggregating microparticles of claim 1 suitable for ocularinjection.
 3. The surface modified solid aggregating microparticles ofclaim 2 suitable for a delivery route selected from the group consistingof intravitreal, intrastromal, intracameral, subtenon, sub-retinal,retrobulbar, peribulbar, suprachoroidal, subchoroidal, conjunctival,subconjunctival, episcleral, posterior juxtascleral, circumcorneal, andtear duct injections.
 4. The surface-modified solid aggregatingmicroparticles of claim 1, wherein the at least one pellet is capable ofsustained drug delivery for at least two months.
 5. The surface-modifiedsolid aggregating microparticles of claim 1, wherein the at least onepellet is capable of sustained drug delivery for at least four months.6. The surface-modified solid aggregating microparticles of claim 1,wherein the at least one pellet is capable of sustained drug deliveryfor at least six months.
 7. The surface-modified solid aggregatingmicroparticles of claim 1, wherein the surface modification is carriedout at a pH between about 14 and about
 12. 8. The surface-modified solidaggregating microparticles of claim 1, wherein the surface modificationis carried out at a pH between about 12 and about
 10. 9. Thesurface-modified solid aggregating microparticles of claim 1, whereinthe surface modification is carried out at a pH between about 10 andabout
 8. 10. The surface-modified solid aggregating microparticles ofclaim 1, wherein the surface modification is carried out at a pH betweenabout 6 and about
 8. 11. The surface-modified solid aggregatingmicroparticles of claim 1, wherein the surface modification is carriedout at a pH between about 6.5 and about 7.5.
 12. The surface-modifiedsolid aggregating microparticles of claim 1, wherein the surfacemodification is carried out at a temperature of less than 16° C.
 13. Thesurface-modified solid aggregating microparticles of claim 1, whereinthe surface modification is carried out at a temperature of less than10° C.
 14. The surface-modified solid aggregating microparticles ofclaim 1, wherein the surface modification is carried out at atemperature of less than 8° C.
 15. The surface-modified solidaggregating microparticles of claim 1, wherein the surface modificationis carried out at a temperature of less than 5° C.
 16. Thesurface-modified solid aggregating microparticles of claim 1, whereinthe therapeutic agent is a tyrosine kinase inhibitor.
 17. Thesurface-modified solid aggregating microparticles of claim 16, whereinthe therapeutic agent is sunitinib or a pharmaceutically acceptable saltthereof.
 18. The surface-modified solid aggregating microparticles ofclaim 17, wherein the therapeutic agent is sunitinib malate.
 19. Thesurface-modified solid aggregating microparticles of claim 1, whereinthe microparticles deliver a therapeutically effective amount of atherapeutic agent selected from dorzolamide, brinzolamide, brimonidine,and timolol.
 20. The surface-modified solid aggregating microparticlesof claim 19, wherein the therapeutic agent is timolol.
 21. Thesurface-modified solid aggregating microparticles of claim 1, whereinthe microparticles have a mean diameter between about 20 and 30 μm. 22.The surface-modified solid aggregating microparticles of claim 1,wherein the microparticles have a mean diameter between about 25 and 35μm.
 23. The surface-modified solid aggregating microparticles of claim1, wherein the microparticles have a mean diameter between about 20 and40 μm.
 24. The surface-modified solid aggregating microparticles ofclaim 1, wherein the microparticles have a mean diameter between about20 and 50 μm.
 25. The surface-modified solid aggregating microparticlesof claim 1, wherein the microparticles have a mean diameter betweenabout 25 and 40 μm.
 26. The surface-modified solid aggregatingmicroparticles of claim 1, wherein the at least one pellet has adiameter of at least 600 μm.
 27. The surface-modified solid aggregatingmicroparticles of claim 1, wherein the at least one pellet has adiameter of at least 1 mm.
 28. The surface-modified solid aggregatingmicroparticles of claim 1, wherein the at least one pellet has adiameter of at least 2 mm.
 29. The surface-modified solid aggregatingmicroparticles of claim 1, wherein the at least one pellet has adiameter of at least 5 mm.
 30. The surface-modified solid aggregatingmicroparticles of claim 1, wherein the at least one pellet is capable ofreleasing not more than about 15 percent by weight of the therapeuticagent in vivo over a 24 hour period.
 31. The surface-modified solidaggregating microparticles of claim 1, wherein the at least one pelletis capable of releasing not more than about 10 percent by weight of thetherapeutic agent in vivo over a 12 hour period.
 32. Thesurface-modified solid aggregating microparticles of claim 1, whereinthe at least one pellet is capable of releasing not more than about 15percent by weight of the therapeutic agent in vivo over a 12 hourperiod.
 33. The surface-modified solid aggregating microparticles ofclaim 1, wherein the microparticles have a drug loading of 1-40 percentby weight.
 34. The surface-modified solid aggregating microparticles ofclaim 1, wherein the microparticles have a drug loading of 5-25 percentby weight.
 35. The surface-modified solid aggregating microparticles ofclaim 1, wherein the microparticles have a drug loading of 5-15 percentby weight.
 36. The surface-modified solid aggregating microparticles ofclaim 1, wherein the microparticles have a drug loading of 0.1-5 percentby weight.
 37. The surface-modified solid aggregating microparticles ofclaim 1, wherein the microparticles comprise poly(lactide-co-glycolide).38. The surface-modified solid aggregating microparticles of claim 1,wherein the microparticles comprise poly(lactide-co-glycolide)covalently linked to polyethylene glycol.
 39. The surface-modified solidaggregating microparticles of claim 1, wherein the microparticlescomprise both poly(lactide-co-glycolide) and poly(lactide-co-glycolide)covalently linked to polyethylene glycol.
 40. The surface-modified solidaggregating microparticles of claim 1, wherein the microparticlescomprise poly(lactic acid).
 41. The surface-modified solid aggregatingmicroparticles of claim 1, wherein the microparticles comprise bothpoly(lactic acid) and poly(lactide-co-glycolide) covalently linked topolyethylene glycol.
 42. The surface-modified solid aggregatingmicroparticles of claim 1, wherein the microparticles comprisepoly(lactide-co-glycolide), poly(lactic acid), andpoly(lactide-co-glycolide) covalently linked to polyethylene glycol. 43.The surface-modified solid aggregating microparticles of claim 1 whereinthe surfactant comprises polyvinyl alcohol.
 44. The surface-modifiedsolid aggregating microparticles of claim 1, wherein the surfacemodification is carried out on wet microparticles.
 45. Thesurface-modified solid aggregating microparticles of claim 1, whereinthe surface treated microparticles are capable of releasing atherapeutic agent over a longer period of time compared to non-surfacetreated microparticles.
 46. The surface-modified solid aggregatingmicroparticles of claim 1, wherein the surface-modified solidaggregating microparticles are more hydrophobic than the microparticlesprior to the surface modification.
 47. The surface-modified solidaggregating microparticles of claim 1, wherein the surface-modifiedsolid aggregating microparticles are less inflammatory than non-surfacetreated microparticles.
 48. An injectable material that comprises themicroparticles of claim 1 in a pharmaceutically acceptable carrier foradministration in vivo.
 49. The injectable material of claim 48, furthercomprising a compound that inhibits aggregation of microparticles priorto injection.
 50. The injectable material of claim 49 wherein thecompound is a sugar.
 51. The injectable material of claim 50 wherein thesugar is mannitol.
 52. The injectable material of claim 48, furthercomprising a viscosity enhancer.
 53. The injectable material of claim52, wherein the viscosity enhancer comprises hyaluronic acid.
 54. Theinjectable material of claim 52, wherein the viscosity enhancercomprises sodium hyaluronate.
 55. The injectable material of claim 48,wherein the injectable material has a range of concentration of thesurface-modified solid aggregating microparticles of about 100-600mg/ml.
 56. The injectable material of claim 48, wherein the injectablematerial has a range of concentration of the surface-modified solidaggregating microparticles of about 500 or less mg/ml.
 57. Theinjectable material of claim 48, wherein the injectable material has arange of concentration of the surface-modified solid aggregatingmicroparticles of about 300 or less mg/ml.
 58. The injectable materialof claim 48, wherein the injectable material has a range ofconcentration of the surface-modified solid aggregating microparticlesof about 200 or less mg/ml.
 59. The injectable material of claim 48,wherein the injectable material has a range of concentration of thesurface-modified solid aggregating microparticles of about 150 or lessmg/ml.